RU2845571C1 - Vaccine based on an adeno-associated type 9 vector encoding spike protein for the prevention of sars-cov-2 coronavirus infection - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: invention relates to molecular biology, biotechnology, and medicine. A genetic construct is described for the expression in human or animal cells of a polynucleotide encoding a full-length spike protein of the SARS-CoV-2 virus containing mutations: partial deletion of the RRAR furine site with the removal of all arginines (R) included in the site, as well as mutations K986P, V987P and D614G. The construct is represented by the adeno-associated AAV9 virus, into which the specified polynucleotide is cloned. A recombinant adeno-associated viral vector AAV9 is described, which provides expression in human or animal cells of a polynucleotide encoding a full-length spike protein of the SARS-CoV-2 virus with a genetic construct. The application of a genetic construct or recombinant adeno-associated viral vector AAV9 for the induction of an immune response against the SARS-CoV-2 virus is presented.

EFFECT: the developed genetic construct and a vaccine based on it expand the arsenal of means for inducing specific immunity to the SARS-CoV-2 virus and can be used to effectively stimulate the humoral and cellular response in order to prevent coronavirus infection caused by SARS-CoV-2.

9 cl, 16 dwg, 6 tbl, 5 ex

Description

Field of technology

The invention relates to molecular biology, biotechnology, medicine and can be used to create a vaccine for the prevention of coronavirus infection caused by the SARS-COV-2 virus.

Prior art

Adeno-associated vectors are the most popular method of gene therapy and delivery of therapeutic antibodies due to their relatively low immunogenicity, high safety profile, broad tropism and tendency to maintain long-term expression of target genes. Adeno-associated vectors are mainly used to treat eye and muscle diseases, however, in recent years, there has been an increasing number of studies using this type of vector as a basis for vaccines for the treatment and prevention of infectious diseases such as HIV, HPV and influenza. A number of studies indicate that vector vaccines based on adeno-associated viruses can induce high levels and long-term persistence of antibodies after a single administration and without the need for an adjuvant [1].

The results of studies have been published demonstrating high immunogenicity and protective efficacy of adeno-associated vectors encoding the RBD fragment of the SARS-CoV-2 virus when administered intramuscularly and intranasally to mice [2]-[4], as well as long-term persistence of antibodies in the blood of immunized animals in a macaque model [5]. Phase 1-2 clinical trials of adeno-associated vector type 5 encoding the RBD fragment of the SARS-CoV-2 virus are underway [6].

Despite the formation of population immunity due to mass vaccination and natural infection, as well as rapid diagnostics and existing treatment approaches, the SARS-CoV-2 virus continues to circulate in all countries of the world, and it can still cause death in people with poor health. Therefore, the search for alternative options for reliable, long-term and safe prevention of coronavirus infection caused by the SARS-COV-2 virus remains an urgent task, and the use of adeno-associated vectors as a basis for creating a vaccine is a promising direction.

Disclosure of invention

The objective of the invention is to develop a vaccine based on adeno-associated virus type 9 to stimulate humoral and cellular immune responses in order to prevent coronavirus infection caused by the SARS-CoV-2 virus.

To solve the problem, a genetic construct was developed for expression in the cells of the target organism of a polynucleotide encoding the full-length spike protein of the SARS-CoV-2 virus, containing a deletion in the furin site RRAR682A, as well as mutations KV986PP and D614G, and characterized by the fact that it is represented by the adeno-associated virus AAV9, into which the polynucleotide is cloned.

In some embodiments, the full-length SARS-CoV-2 spike protein comprises additional mutations specific to a particular SARS-CoV-2 variant. In particular embodiments, the SARS-CoV-2 variant is, but is not limited to, an Alpha variant, a Beta variant, a Delta variant, or an Omicron variant.

In some particular embodiments of the invention, the full-length spike protein of the SARS-CoV-2 virus has the sequence of SEQ ID NO: 2 or SEQ ID NO: 3 or SEQ ID NO: 6.

In some particular embodiments, the polynucleotide encoding the full-length spike protein of the SARS-CoV-2 virus is represented by the sequence SEQ ID NO: 7 or SEQ ID NO: 8 or SEQ ID NO: 11.

The solution to the problem is also achieved by developing a recombinant adeno-associated viral vector AAV9, capable of expressing in human or animal cells a polynucleotide encoding the full-length spike protein of the SARS-CoV-2 virus, as described above.

In some embodiments of the invention, the recombinant adeno-associated viral vector AAV9 is represented by the sequence SEQ ID NO: 12 or SEQ ID NO: 13 or SEQ ID NO: 16.

The stated task is also solved by developing a vaccine for the prevention of coronavirus infection in humans or animals, containing at least one genetic construct, as described above, as an active agent, in an effective amount, as well as a pharmaceutically acceptable carrier and a buffer solution.

The problem is also solved by using the developed recombinant adeno-associated viral vector AAV9 (developed vaccine) to induce (activate) an immune response against the SARS-CoV-2 virus and use it as a vaccine preparation for immunization against various variants of SARS-CoV-2 virus strains, including new strains.

As part of the solution to the problem, a method for preventing SARS-CoV-2 coronavirus infection is also proposed by intramuscular administration of the developed vaccine to a subject in need of protection from the infection in an effective dose.

As a result of implementing the invention, the following technical results are achieved:

- the developed genetic construct and the vaccine based on it expand the arsenal of means for inducing specific immunity to the SARS-CoV-2 virus and can be used to effectively stimulate the humoral and cellular response in order to prevent coronavirus infection caused by SARS-CoV-2;

- due to the introduced mutations aimed at eliminating infectivity and imparting stability to the expressed antigen, ensuring the safety and effectiveness of the proposed design, as well as due to the possibility of using designs containing additional mutations inherent in various variants of the SARS-CoV-2 virus, the developed genetic design is a universal basis for developing vaccines for the purpose of immunization against various variants of SARS-CoV-2 virus strains, including new strains;

- the use of the adeno-associated vector type 9 (AAV9) in the design of the developed vaccine, together with other features of the genetic design, ensures the duration of expression of the design for more than 24 weeks after one injection, high efficiency of transduction in muscles, safety ensured by local production of the antigen at the injection site, scalability of production, which allows the use of the developed genetic design for the production of a single-component vaccine for intramuscular administration.

Brief description of the drawings

Fig.1 A - D - Graphic representation of the developed designs.

Fig.2 A - E - Genetic maps of the developed plasmids pAAV-SARS1 - pAAV-SARS5 and pAAV-Luc-control (pAAV-Luc).

Fig. 3 - Genetic maps of the developed plasmids pAAV-SARS1 - pAAV-SARS5 and pAAV-Luc-control based on the results of restriction: a schematic arrangement of plasmid fragments after analytical restriction (Theoretical map) and the result of the restriction hydrolysis of the obtained plasmids (Restriction result) are shown. The map was compiled in the Benchling program, the restriction fragment length diagram was compiled in the SerialCloner program.

Fig. 4 - Results of dot blot analysis of transduced cell lysates. Numbers indicate: 1-5 - cells transduced with rAAV9-SARS1 - rAAV9-SARS5 vectors, respectively; 6 - negative control (non-transduced cells); 7-9 - positive control (cells infected with B.1 coronavirus, B.1.617.3 Delta coronavirus, or AY.122 Russian Delta coronavirus, respectively).

Fig. 5 - Dynamics of mice weight after immunization with experimental adeno-associated vectors. The graph shows the change in mice body weight in % relative to the initial value, average values for the group and standard deviation are given (see Example 5).

Fig. 6 - Dynamics of neutralizing antibodies to the Delta variant of the SARS-CoV-2 virus in the sera of immunized mice (see Example 5). The dots represent individual values for each animal, the columns represent the geometric mean titers by group.

Fig. 7 - Dynamics of virus-specific IgG antibodies to the Delta variant of the SARS-CoV-2 virus in the sera of immunized mice (see Example 5). The dots represent individual values for each animal, the columns represent the geometric mean titers by group.

Fig. 8 - Sections of the lung (A), heart (B), liver (C) and kidney (D) of mice 3.11 and 3.12 (see Example 6). Staining: hematoxylin and eosin. Magnification ×100.

Fig. 9 - Cross-section of skeletal muscles of mice 3.11 (A, B) and 3.12 (B), obtained 24 weeks after the introduction of the vector vaccine (see Example 6). Staining: hematoxylin and eosin. A, B - magnification ×100; B - magnification ×400.

Fig. 10. - Immunofluorescence detection of S-protein: (A) - positive control, (B) - negative control, (C) - experimental group (see Example 6). S-protein is marked in red, autofluor - in green, the nucleus is stained with DAPI - in blue.

Fig.11 - Dynamics of mice weight after immunization with experimental adeno-associated vectors. The graph shows the change in mice body weight in % relative to the initial weight, average values for the group and standard deviation are given (see Example 7).

Fig. 12 - Dynamics of neutralizing antibodies to the Beta variant of the SARS-CoV-2 virus in the sera of mice immunized with rAAV9-SARS2, rAAV9-SARS3 or rAAV9-SARS4 vaccines (see Example 7). The dots represent individual values for each animal, the columns represent the geometric mean titers by groups. **** p < 0.0001, statistically significant change in titer between 4 and 12 weeks (ANOVA, Tukey's pairwise test).

Fig. 13 - Dynamics of virus-specific IgG antibodies to the Beta variant coronavirus in the sera of immunized mice (see Example 7). The dots represent individual values for each animal, the columns represent geometric mean titers by groups. **** p < 0.0001, statistically significant change in titer between 4 and 12 weeks, ns - not significant (ANOVA, Tukey's pairwise test).

Fig. 14 - Dynamics of virus-specific IgG antibodies to the AAV9 capsid protein in the sera of immunized mice (see Example 7). The dots represent individual values for each animal, the columns represent geometric mean titers by groups. ** p < 0.01, statistically significant change in titer between 4 and 12 weeks, ns - not significant (ANOVA, Tukey's pairwise test).

Fig. 15 - Cumulative antigen-specific production of cytokines by CD4+ and CD8+ effector memory T cells in response to immunization with the studied drugs. The graphs show individual values of the cumulative proportion of cytokine-producing T lymphocytes from the total number of effector CD4+ (A) and CD8+ (B) T cells (CD4/8+CD44+CD62L-) upon stimulation with PepTivator S. The groups were compared using one-way ANOVA with subsequent comparison of the study groups with the group of naive animals using Dunnett's test, * p < 0.05, ** p <0.001; *** p <0.0001; ns p > 0.05.

Fig. 16 - Subpopulation composition of cytokine-producing CD4+ and CD8+ Tem in mouse spleens in response to immunization with the studied preparations. The graphs show data reflecting the percentage of various subpopulations of cytokine-producing CD4+ and CD8+ cells in the total CD4/8+CD44+CD62L-Tem population. Groups were compared using ANOVA followed by pairwise comparison using the Tukey test. *: p<0.05, **: p<0.01, ***: p<0.001.

Terms and definitions

The following terms and definitions are used in this document unless otherwise explicitly stated. References to techniques used in describing this invention refer to well-known techniques, including variations of these techniques and their replacement by equivalent techniques known to those skilled in the art.

In the documents of this invention, the terms "includes", "including" , etc., as well as "contains", "comprising" , etc. are interpreted to mean "includes, among other things" (or "contains, among other things"). These terms are not intended to be interpreted as "consists only of".

The term "and/or" means one, more, or all of the listed elements.

The term "optional" or "optional " or "optionally" as used herein means that the subsequently described event or circumstance may, but need not, occur and that the description includes instances in which the event or circumstance occurs and instances in which it does not occur.

The terms “target organism” (“subject”) and “patient” refer to a human or animal (mammal) exposed to infection with the SARS-CoV-2 virus.

The terms "treatment", "therapy" mean cure, slowing, stopping, or reversing the progression of a disease or disorder (alleviating the severity of the disease). In this document, "treatment" also means alleviating the symptoms associated with the disease or disorder.

The term "prophylaxis", "prevention" covers the elimination of risk factors, as well as prophylactic treatment of subclinical stages of the disease in a patient, aimed at reducing the likelihood of the occurrence of clinical stages of the disease. Primary prophylaxis is defined as prophylactic administration of the composition to patients whose clinical stage of the disease has not yet occurred. Secondary prophylaxis is the prevention of the recurrence of the same or a similar clinical state of the disease.

The "vaccine"("vaccinecomposition") according to the invention comprises a genetic construct developed within the framework of the invention as an active agent in an effective amount, as well as a physiologically acceptable carrier and a buffer solution. Pharmaceutically acceptable carriers and buffer solutions are well known in the art and include those described in various textbooks, such as, for example, Remington's Pharmaceutical Sciences.

Unless otherwise defined, technical and scientific terms in this application have the standard meanings generally accepted in the scientific and technical literature.

The numbers of the nucleotide and amino acid sequences referred to in this application correspond to the number in the Sequence Listing (SEQ ID NO) according to Standard ST.26, which is part of this description of the invention. In the event of discrepancies in the structure of the sequences between the mention in the text of the description and the corresponding sequence in the Sequence Listing according to Standard ST.26, the data given in the text of the description shall prevail.

Detailed description of the invention and its implementation

As part of the implementation of the invention, recombinant adeno-associated vectors have been developed that are capable of producing antigens in human and mammalian cells, in response to which protective immunity against the SARS-CoV-2 virus is formed, and which can thus be used as a vector vaccine for the prevention of SARS-CoV-2 coronavirus infection. Due to the fact that the active principle of the vaccine is precisely the genetic construct with which the antigen is synthesized, as well as due to the features of the developed genetic construct, the use of the developed vaccine makes it possible to obtain an immune response profile that includes both a cytotoxic and an antibody immune response, with the formation of immunological memory.

To implement the present invention, antigen construct variants were developed at the first stage. The S-protein (Spike, spike protein) of the SARS-CoV-2 virus was taken as the basis for the construct. At the same time, in order to increase the stability of the expressed antigen, targeted mutations were introduced into its construct. As is known, the S-protein of the SARS-CoV-2 virus contains a cleavage site (the so-called furin site), recognized by the cellular protease furin, located at the junction of the S1 and S2 subunits and represented by RRAR amino acids (corresponding to positions 682-685 of the sequence SEQ ID NO: 1 of the S-protein of the SARS-CoV-2 virus of the Wuhan strain). Cleavage of the S-protein by furin activates the virus during its interaction with the cell. It has been established that the absence of a cleavage site significantly limits the spread of the virus in the body, therefore, many variants of targeted mutations of this site are known, the purpose of which is to inactivate it for cleavage by furin (see, for example, [7], [8]). Most of such targeted mutations represent a partial or complete replacement of RRAR amino acids, some variants also offer a complete deletion of the cleavage site. Within the framework of the present invention, it is proposed to use a partial deletion of the cleavage site, namely, the removal of all arginines included in the site. Thus, the construct according to the invention contains a mutation in the furin site RRAR682A. Due to the presence of such a targeted mutation, the stability of the antigen expressed as a result of immunization increases, and important antigenic determinants for the formation of humoral immunity are preserved.

In order to provide stability to the antigen, a targeted mutation was also introduced into its design, consisting of replacing amino acids at positions 986 and 987 of SEQ ID NO: 1 with proline - KV986PP (the so-called double mutation to proline), widely used in the S-protein stabilization strategy in vaccine development (see, for example, [9]). Another targeted mutation is the D614G mutation, which is natural for many strains [10], but provides additional stability to the antigen. Due to the presence of several mutations in the developed design that provide stability, the antigen expressed as a result of vaccination has increased stability.

In the context of vaccine development, both full-length engineered S proteins of different SARS-CoV-2 virus strains (i.e., the protein consisting of 1270 amino acid residues) and truncated S protein variants, such as only the S1 subunit (amino acid residues 14-685) and the receptor-binding domain, RBD protein (amino acid residues 319-541), were studied. A graphical representation of the engineered antigen constructs is shown in Fig. 1.

As the experiments conducted have shown (see details in the Examples section), although the use of the S1 subunit and the receptor-binding domain of the RBD has shown its effectiveness in previously developed vaccines of a different type against the SARS-CoV-2 virus, these constructs unexpectedly turned out to be insufficiently effective in the vector vaccines under development.

As is known, different strains (variants) of the SARS-CoV-2 virus differ from each other by the presence of mutations, including in the structure of the S protein. A person skilled in the art understands that different strains (variants) of the SARS-CoV-2 virus can be used in the production of the vaccine according to the invention, which will determine the effectiveness of vaccination against a specific strain (variant), but does not affect the approach to constructing a genetic construct based on the adeno-associated virus according to the invention. During the development of the vaccine, constructs containing mutations characteristic of different variants of the SARS-CoV-2 virus were analyzed, such as the Wuhan strain, the Beta variant, and the Delta variant. The analysis showed that the effectiveness of immunization does not depend on the presence of mutations in the construct inherent in a specific variant of the virus. Thus, the proposed design approach can be used to develop a vaccine against various strains (variants) of the SARS-CoV-2 virus, such as known variants (Alpha variant, Beta variant, Delta variant, Omicron variant, etc.) or potentially new strains (variants) of the SARS-CoV-2 virus.

The adeno-associated virus 9 (AAV9) was used as a vector in the development of the vaccine; it has a high tropism for muscles and good scalability in production [11].

Recombinant adeno-associated virus (rAAV) products are produced using transient cell transfection methods. These methods provide the highest yields and also produce a safe and effective rAAV product expressing the transgene [12]. This method involves transfecting HEK293 cells with two or three plasmids (forming a so-called plasmid system): the first encodes the target gene, the second carries the AAV rep and cap genes, and the third contains adeno- or herpesvirus helper genes. AAV production can be accomplished on a large scale using a number of manufacturing methods that have proven themselves in the industry to produce dozens of products currently in clinical development and available commercially [13], [14].

In order to implement the present invention, plasmid systems necessary for assembling rAAV according to the developed variants of antigen constructs were obtained, including plasmids: a plasmid encoding a target gene (selected from pAAV-SARS1 - pAAV-SARS5), a pAAV-RC2/9 plasmid (carrying the rep gene of serotype AAV2 and the cap gene of serotype AAV9) and a pHelper plasmid. The necessary recombinant plasmids were obtained in preparative quantities by transforming E. coli cells of the Stbl3 strain. It is known to specialists in this field that any other plasmid systems containing a nucleotide sequence encoding an antigen according to the invention can be used to obtain a recombinant adeno-associated viral vector.

Next, to obtain the rAAB9 vector expressing the SARS-CoV-2 virus antigen of the invention, the obtained plasmids were transfected into HEK293T cells, in which the production and assembly of the recombinant adeno-associated virus carrying the mutated gene of the full-length S protein of the SARS-CoV-2 virus took place, according to various embodiments of the design of the invention. The purification of the recombinant adeno-associated virus was carried out using tangential filtration and affinity chromatography. Those skilled in the art will understand that other eukaryotic expression systems can be used for the purpose of implementing the present invention for the production and assembly of the recombinant adeno-associated viral vector. If necessary, the production method further includes isolation and purification by any method known in the art.

Experts also understand that the obtained genetic constructs include, in addition to the polynucleotide encoding the SARS-CoV-2 virus antigen, all the necessary elements that make it possible to implement the specified purpose - to express the polynucleotide in target cells.

Detailed information on the production of a vaccine based on the adeno-associated vector of serotype 9 encoding the S protein for the prevention of coronavirus infection SARS-CoV-2 is given in the following detailed examples.

It should be understood that the examples given in the application materials are not limiting and are provided only to illustrate the present invention.

Example 1. Design of an antigen construct

The following design variants were constructed as antigens for the vaccine:

1) The full-length S protein of the Wuhan strain containing a deletion in the furin site of RRAR682A and mutations to proline in the amino acids of KV986PP, as well as the D614G mutation (see SEQ ID NO: 2 and Fig. 1A);

2) The full-length S protein containing a deletion in the furin site RRAR682A and mutations to proline in the amino acids KV986PP, as well as the D614G mutation and two additional mutations in the RBD domain, characteristic of the Beta variant of the SARS-CoV-2 virus (see SEQ ID NO: 3 and Fig. 1B);

3) The S1 subunit of the S protein containing the D614G mutation, as well as two additional mutations in the RBD domain, characteristic of the Beta variant of the SARS-CoV-2 virus (see SEQ ID NO: 4 and Fig. 1B);

4) RBD protein containing the D614G mutation, as well as two additional mutations in the RBD domain characteristic of the Beta variant (see SEQ ID NO: 5 and Fig. 1G);

5) Full-length S protein containing a deletion in the furin site RRAR682A and proline mutations in amino acids KV986PP, as well as the D614G mutation, and three additional mutations in the RBD domain characteristic of the Delta variant of the SARS-CoV-2 virus (K417N, L452R, T478K) (see SEQ ID NO: 6 and Fig. 1D).

After the development of antigen constructs, the nucleotide sequences encoding them were synthesized (SEQ ID NO: 7 - SEQ ID NO: 11, respectively), which were subsequently used to obtain target plasmids.

Example 2. Obtaining plasmid constructs

The target plasmids pAAV-SARS1 to pAAV-SARS5 and the control plasmid pAAV-Luc-control were obtained by ligation-free cloning. For this purpose, primers with overlapping sequences were used, as indicated in Table 1.

Table 1. Primer sequences for cloning

Name of the design Primer name Primer sequence 5'-3' SEQ ID NO: pAAV-SARS1-
pAAV-SARS4 Var1F agacgctagactagcatgcgactcttcgcgatgtacgggccag 17 Var1R caattgttgttgttttgctgtccataaaaccgcccagtagaag 18 VecVar1F tatggacagcaaaacaacaacaattggatctgcgctgattttgt 19 VecVar1R gtcgcatgctagtctagcgtctaaacgcgt 20 pAAV-SARS5 Var5F ATGGACCTGGAAGGCAAGCAGGg 21 Var5VecR CCCTGCTTGCCTTCCAGGTCCATC 22 pAAV-Luc-control LucF ctgGCTAGCCGCCACCatggaagacgccaaaaacataaagaaagg 23 LucVecR GGTGGCGGCTAGCcagcttg 24 LucVecF tgtaagggcccgtttaaacccg 25 LucR cgggtttaaacgggcccttacacggcgatctttccgcc 26

The pAAV-CMV construct was used as a vector for cloning the coding sequences. Inserts were amplified from pVax1-Var1-5 plasmids. Maps of the resulting pAAV-SARS1 - pAAV-SARS5 plasmids and the control pAAV-Luc-control plasmid are shown in Fig. 2.

The obtained constructs were transfected into E. coli cells, strain Stbl3. For transfection on Petri dishes (150 ^cm2 ), a seeding density of 50,000 cells/ ^cm2 was used. Cell growth on dishes before transfection took 48 hours. The composition of the cultivation medium: DMEM with the addition of 4.5 g/l glucose, 10% fetal calf serum, streptomycin, penicillin and replaceable amino acids. Three hours before adding the transfection mixture, the medium was completely replaced with a depleted one. The morphology and confluency of the cell culture were monitored on a control Petri dish. At the time of transfection, the cell culture density did not exceed 75-80%. For transfection on a dish, the volume of the medium for the time of transfection was 20 ml. Next, the selection of transformant clones was performed on a selective nutrient medium and the selected clone was cultivated to obtain preparative quantities of plasmids carrying the target genes. After lysing the cells, the plasmids were isolated using chromatographic purification.

To check the obtained plasmids, restriction hydrolysis was performed using specific restriction endonucleases. The genetic map of the obtained plasmids - a schematic arrangement of plasmid fragments after analytical restriction and the result of restriction hydrolysis of the obtained plasmids - is shown in Fig. 3. The map was compiled in the Benchling program, the restriction fragment length scheme - in the SerialCloner program. As the analysis showed, all fragments obtained as a result of restriction hydrolysis fully correspond to the theoretically calculated ones. Plasmids were also checked by Sanger sequencing.

Example 3. rAAV production in HEK293T cell culture

To obtain the genetic constructs required for the assembly and production of the recombinant adeno-associated viral vector, the transient transfection method of HEK293T cells was used. For this purpose, plasmid systems required for the assembly of rAAV according to the developed variants of the antigen constructs were used, and including the plasmids: (1) a plasmid carrying a target gene having a sequence selected from SEQ ID NO: 7 - SEQ ID NO: 11 (respectively, a plasmid selected from pAAV-SARS1 - pAAV-SARS5) (or pAAV-Luc-control - for the control vector), obtained as described in Example 2; (2) plasmid pAAV-RC2/9 (carrying the rep gene of serotype AAV2 and the cap gene of serotype AAV9); and (3) plasmid pHelper.

For transient transfection of HEK293T cells, a mixture of three plasmids was prepared in DMEM medium in the following amounts: (1) target (0.056 μg/cm ² ), (2) pAAV-RC2/9 (0.055 μg/cm ² ), and (3) pHelper (0.089 μg/cm ² ) in a total volume of 200 ml of transfection medium (0.005 ml/cm ² ) to obtain a “DNA/DMEM” mixture. Linear polyethyleneimine (LPEI) 25 kDa was used as a transfection reagent. The “DNA/DMEM” and “LPEI/DMEM” mixtures were mixed under sterile conditions and thoroughly mixed for 1 minute. The prepared mixture was incubated for 15 minutes at room temperature. For transfection on Petri dishes, the volume of the “LPEI/DMEM” mixture and the “DNA/DMEM” mixture was 1.5 ml for each dish, the premix was prepared in 50 ml tubes for 10 dishes. For transfection on Petri dishes, the volume of the medium did not change, 3 ml of the transfection mixture was added dropwise to 20 ml of the medium and distributed by shaking.

72 hours after transfection, 0.5% Triton X-100 was added directly to the cell suspension. It was collected aseptically in a glass bottle. The lysate was centrifuged in 0.25 L beakers at 3000 g, 10 min, +4°C. The clarified lysate (semi-finished product) was collected, samples for quantitative assessment (50 μl) were taken and frozen at -20°C. Viral production was monitored by assessing the rAAV virus titer in the lysate using quantitative real-time PCR. A result of more than 5E+11 genomic copies in 1 ml was considered acceptable.

Viral particles were isolated by iodixanol gradient ultracentrifugation. For this purpose, the following were layered in succession into Ultra-clear 16x76mm tubes (Beckman Coulter) for a 70Ti rotor: 2 ml “60%” fraction (58.6% iodixanol), 2 ml “40%” fraction (40% iodixanol), 2.75 ml “25%” fraction (25% iodixanol) and 4.5 ml of the resulting viral preparation. Centrifugation was performed in an iodixanol density gradient at 350,000 g and +18°C for 1 hour in a Beckman Optima-XPN or Optima-XE ultracentrifuge. Fractions 60%+½ 40% were then collected.

The sample was dialyzed in 100 kDa dialysis bags (Spectrum, #0867140) against 1xPBS/350 mM NaCl/0.001% Pluronic F-68 buffer at +4°C. The buffer was changed 2 to 3 times over 14-18 h. It was ensured that the volume of the dialysis buffer was at least 200 times greater than the sample volume. Concentration was performed in 100 kDa centrifugal concentrators to a calculated concentration of 3E+13 genomic copies in 1 ml. The resulting viral preparation was sterilized through spin columns with a pore size of 0.22 μm and frozen in a buffer for storing viral preparations (1x PBS, 350 mM NaCl, 0.001% Pluronic F-68) in 0.5 ml Eppendorf tubes. The concentration of viruses was determined by quantitative real-time PCR using probes and primers to inverted repeats. As a result of the experiment, recombinant adeno-associated viral vectors rAAV9-SARS1 - rAAV9-SARS5, carrying target genes and having sequences presented by SEQ ID NO: 12 - SEQ ID NO: 16, respectively, suitable for in vitro and in vivo tests, with a concentration of 1x10 ¹² genomic copies in 1 ml of each, as well as a control luciferase vector rAAV9-Luc, having a sequence presented by SEQ ID NO: 27, were collected and purified.

Example 4. Study of expression of target proteins (antigens) by experimental adeno-associated vectors during cell transduction in vitro

Expression of target proteins, including different constructs of the SARS-CoV-2 S protein, was assessed by transducing HEK293T cells with experimental adeno-associated vectors rAAV9-SARS1 - rAAV9-SARS5. After transduction, the cells were lysed, the cell lysates were applied to the membrane and then examined using the staining method with RBD-specific antibodies to the RBD domain of the SARS-CoV-2 S protein - XR-06, XR-25 (Hema LLC) or a mixture of them. Untransduced HEK293T cells were used as a negative control, and HEK293T cells infected with the B.1 coronavirus, the Delta B.1.617.3 coronavirus, or the Russian Delta AY.122 coronavirus were used as a positive control.

As shown by the results of dot blot analysis, all variants of experimental adeno-associated vectors expressed the RBD fragment, while the expression level varied and was highest in the rAAV9-SARS4 and rAAV9-SARS3 variants (see Fig. 4).

Example 5. Study of the expression of the target Spike protein of the SARS-CoV-2 Delta variant by experimental adeno-associated vectors and their immunogenicity in vivo

The study was conducted on BALB/c mice, females, 6-8 weeks old. The mice were obtained from the Stolbovaya nursery of the Federal State Budgetary Scientific Institution "National Center for Biomedical Technologies" of the Federal Medical and Biological Agency of Russia, Moscow Region. All experiments involving laboratory animals were approved by the Bioethics Committee of the A.A. Smorodintsev Research Institute of Influenza of the Ministry of Health of the Russian Federation, Protocol No. 55 dated April 4, 2022.

After the end of quarantine, the animals (45 animals in total) were vaccinated by intramuscularly injecting 100 μl of the preparation into the hind paw. Groups 1-3 were administered rAAV Delta variant of the SARS-CoV-2 virus (rAAV9-SARS5) at doses of 10 ⁹ genomic copies (GC), 10 ¹⁰ GC and 0.75*10 ¹¹ GC, respectively. The control group (No. 4) was administered the luciferase vector (rAAV9-Luc; Luc control) at a dose of 0.75*10 ¹¹ GC. Group No. 5 consisted of intact animals (see Table 2).

The body weight and condition of the animals were monitored for 2 weeks, weighing them once every 2-3 days. After 4, 12 and 24 weeks, all animals (alive at the time of the experiment) had their blood serum collected during their lifetime to detect virus-specific IgG antibodies and neutralizing antibodies.

After 4, 12 and 24 weeks, organs and tissues (serum, lungs, liver, kidneys, muscles, heart) were collected from five animals from groups 3 (rAAV9-SARS5) and 4 (rAAV9-Luc), and after 4 and 24 weeks from five animals from group 5 (intact) to detect expression of target proteins. Organs from 2/5 animals after extraction were placed in 10% formalin for histological analysis.

Table 2. In vivo study design

№ gr. Preparation Dose Number of mice Observations and manipulations by day 0 1-2 weeks Week 4 Week 12 24th week 1 rAAV9-SARS5 10 ⁹ GK 5 B, I (5) IN C (5) C (5) C (5) 2 rAAV9-SARS5 10 ¹⁰ GK 5 B, I (5) IN C (5) C (5) C (5) 3 rAAV9-SARS5 0.75*10 ¹¹ GK 15 B, I (15) IN C (15), O (3+2) C (10), O (3+2) C (5), O (3+2) 4 rAAV9-Luc 0.75*10 ¹¹ GK 15 B, I (15) IN C (15), O (3+2) C(10), O(3+2) C (5), O (3+2) 5 Intact - 10 IN IN C (10), O (3+2) - C (5), O (3+2)

Note: GC - genomic copies, and - immunization, B - weighing (once every 2-3 days, 2 weeks), C - serum sampling, O - collection of organs and tissues for PCR (3 animals) and IHC (2 animals). The number of animals is given in brackets.

During the 2 weeks after immunization, the weight of the mice steadily increased (see Fig. 5). The exception was a short-term drop in the weight of the mice in group 3 (rAAV9-SARS5, maximum dose), observed on day 13. However, in most mice of this group, the weight was restored by day 14 of observation.

The presence of virus-neutralizing antibodies was determined in animal sera by the classical RMN (microneutralization reaction) method [15], [16] using the SARS-CoV-2 Delta virus (hCoV-19/Russia/SPE-RII-32759V3/2021 strain, line B.1.617.3, GISAID database number: EPI_ISL_8112510) at a dose of 100 TCID50 per well (TCID50 is a tissue cytopathic infectious dose at which 50% damage occurs). The neutralizing titer was considered to be the highest dilution of the serum at which there were no signs of CPE (cytopathic effect) in the well. The study of the antibody response in animals vaccinated with rAAV9-SARS5 showed that mice formed virus-neutralizing antibodies already 4 weeks after immunization. Antibody titers increase by the 12th week after immunization and have a clear dose dependence (see Fig. 6). As can be seen from Fig. 6, in the maximum dose group (0.75*10 ¹¹ GC/mouse), 4 weeks after immunization, the formation of virus-neutralizing antibodies was observed in 85% of animals, while at lower immunizing doses, virus-neutralizing antibodies were formed in only 10-20% of animals after 4 weeks. After 12 weeks after immunization, the formation of virus-neutralizing antibodies was observed in 100% of animals from the maximum and medium dose groups (10 ¹⁰ GC/mouse), while in the lowest dose group (10 ⁹ GC/mouse), virus-neutralizing antibodies were detected in 4/5 animals after 12 weeks. At 24 weeks post-immunization, virus-neutralizing antibodies were observed in 100% of vaccinated animals with a clear dose-dependence of the geometric mean titer (see Table 3).

Table 3. Geometric mean values of antibody titers in groups of immunized animals

Group 1 Group 2 Group 3 Group 4 Group 5 Dot 10 ⁹ GK/mouse 10 ¹⁰ GK/mouse 0.75x10 ¹¹ GK/mouse Luc-control Intact Formation of virus-neutralizing antibodies 4 weeks 6.6* 8.7 25* 5 5 12 weeks 52.8** 279** 557** 5 24 weeks 45.9*** 80 422*** 5 5 IFA 4 weeks 151**** 696 1600**** 66 76 12 weeks 1838***** 6400***** 13719**** 76 24 weeks 919**** 3200****** 12800****** 115 115

Note: Asterisks indicate statistically significant differences between the corresponding groups of animals that received adeno-associated vectors at different doses: *p=0.0109; **p=0.0003; ***p=0.0074; ****p<0.0001; *****p=0.0312; ******p=0.264

The presence of virus-specific IgG antibodies was determined by ELISA with samples of whole animal serum. The antigen used was a preparation of the inactivated purified in a sucrose gradient virus variant Delta SARS-CoV-2.

Similar dynamics as in the case of virus-neutralizing antibodies were observed for virus-specific IgG antibodies detected in ELISA with the whole inactivated SARS-CoV-2 virus as an antigen (see Fig. 7). Since, unlike the neutralization reaction, ELISA has greater sensitivity, antibody titers and the percentage of animals with virus-specific antibodies according to ELISA data were higher. Thus, already 12 weeks after immunization, 100% of animals developed virus-specific IgG antibodies.

At 24 weeks after immunization, a decrease in the geometric mean antibody titer was observed in immunized animals compared to the 12-week point, both according to the neutralization reaction and according to the ELISA data.

Example 6. Morphological and immunofluorescent characteristics of tissues and organs in mice after intramuscular administration of a vector vaccine encoding the Spike protein of the SARS-CoV-2 virus

The material used for the work was fixed in 10% buffered formalin, obtained in the experiment described in Example 5: tissues of animals of groups No. 3 (vaccination with rAAV9-SARS5 at a dose of 0.75x10 ¹¹ GC), No. 4 (immunization with rAAV9-Luc) and No. 5 (intact mice) were studied. The prepared material was distributed among the groups presented in Table 4.

Table 4. Distribution of animal samples by groups

Groups No. Term Organs rAAV9-SARS5 3.1 4 weeks Skeletal muscles, heart, lungs, liver, kidneys 3.2 4 weeks Skeletal muscles, heart, lungs, liver, kidneys 3.7 12 weeks Skeletal muscles, heart, lungs, liver, kidneys 3.8 12 weeks Skeletal muscles, heart, lungs, liver, kidneys 3.11 24 weeks Skeletal muscles, heart, lungs, liver, kidneys 3.12 24 weeks Skeletal muscles, heart, lungs, liver, kidneys rAAV9-Luc 4.1 4 weeks Skeletal muscles, heart, lungs, liver, kidneys 4.2 4 weeks Skeletal muscles, heart, lungs, liver, kidneys 4.7 12 weeks Skeletal muscles, heart, lungs, liver, kidneys 4.8 12 weeks Skeletal muscles, heart, lungs, liver, kidneys 4.11 24 weeks Skeletal muscles, heart, lungs, liver, kidneys 4.12 24 weeks Skeletal muscles, heart, lungs, liver, kidneys Intact 5.1 4 weeks Skeletal muscles, heart, lungs, liver, kidneys 5.2 4 weeks Skeletal muscles, heart, lungs, liver, kidneys 5.6 24 weeks Skeletal muscles, heart, lungs, liver, kidneys 5.7 24 weeks Skeletal muscles, heart, lungs, liver, kidneys

Note: No. is the number assigned to a specific animal.

The fixed material was placed in histological cassettes and washed for 30 minutes in running water. Then the cassettes were processed in a battery of alcohols and paraffins in accordance with the manual wiring protocol, after which the material was poured into paraffin blocks and microtomed. Microtomy was performed on a ThermoFisher HM430 sled microtome to obtain sections 3-5 μm thick. The sections were transferred to glass slides (Biovitrum, Russia) with adhesive liquid (Biovitrum, Russia) previously applied to the surface. Sections on glass slides were dried in a thermostat at 37°C for 12 hours, after which they were deparaffinized, rehydrated, stained in a solution of hematoxylin (Biovitrum, Russia), eosin (Biovitrum, Russia), dehydrated, cleared, and embedded in a polystyrene-based mounting medium under a cover glass.

Autopsy material (lungs) of patients who died from verified SARS-CoV-2 infection was used as a control. The preparations were made in a similar manner, as well as using the classical immunohistochemical method with staining of the preparations with hematoxylin and study in a "bright field".

As shown by the histochemical examination of the internal organs of the animals included in the experiment, no pathological changes were found in the lungs, heart, liver and kidneys in all groups of animals and at all time points from 4 to 24 weeks. Fig. 8 shows sections of the lung (A), heart (B), liver (C), kidney (D) of mice 3.11 and 3.12 (animals from the experimental group, 24 weeks after the introduction of the viral vector). No pathological changes were found.

Histochemical examination of the vaccine injection site (skeletal muscles) was performed on the material represented by the cross-section of several skeletal muscles. On the 4th week of the study, using hematoxylin and eosin staining on the preparation, one animal was found to have both muscles with multiple foci of endomysial infiltration and intact muscles without signs of pathological processes. Foci of endomysial infiltration are clusters of histiocytes and lymphocytes around a viable muscle fiber with an internalized nucleus. Skeletal muscles with signs of endomysial inflammation are also characterized by the presence of a pronounced regenerative pattern - a high proportion of muscle fibers with internalized nuclei (70-90%), a rounded shape of muscle fibers and endomysial lipomatosis, which significantly distinguishes them from the intact skeletal muscles of the same animal.

At week 12, both muscles with multiple foci of endomysial infiltration and intact muscles were also determined in preparations of mouse 3.7. In this case, a smaller number of foci of endomysial infiltration, a decrease in their severity, a decrease in the number of muscle fibers with an internalized nucleus, and a decrease in the number of foci of endomysial lipomatosis are characteristic. In mouse 3.8, foci of inflammatory infiltration were not detected.

At the 24th week, in the skeletal muscle preparations of mice 3.11 and 3.12, there are no signs observed at earlier stages; residual phenomena are represented by single adipocytes in the endo- and perimysium (see Fig. 9A - the only inflammation focus located around the arteriole, detected in mouse 3.11), as well as one periarteriolar lymphohistocyte cuff in the preparation of mouse 3.11 (see Fig. 9B - skeletal muscle of mouse 3.12 without pathological changes and Fig. 9C - periarteriolar cuff of lymphocytes and histiocytes).

In the skeletal muscle preparations of mice group No. 4 (immunization with rAAV9-Luc) and No. 5 (intact mice), pathological changes were not detected.

Thus, in the muscles of mice (3.1, 3.2, 3.7, 3.8, 3.11, 3.12) that were administered the vector vaccine, multiple foci of endomysial inflammation, damage and a hyperregenerative pattern of reactive changes in the muscle are determined - the prevalence of fibers with a central location of the nucleus at the early stages of the experiment (a reaction to damage during acute inflammation), endomysial lipomatosis - replacement of necrotic muscle fibers. The severity of the listed signs decreases with increasing time from the moment of administration of the vector vaccine.

Signs of local damage and inflammation were not detected in the groups with the introduction of a viral vector with luciferase (No. 4) and the intact control group (No. 5), which indicates a relationship between the detected reactive changes and the introduction of a vector vaccine and indicates that the introduction of the vaccine leads to the synthesis of an immunogen protein that causes local manifestations of a transient inflammatory reaction.

The inflammatory reaction was determined only in the area of vector vaccine administration - in skeletal muscles, leaving adjacent muscle groups intact. In all animals of all groups, no pathological changes in internal organs were detected, which allows us to state the absence of dissemination of the viral vector beyond the administration area and the development of any significant and persistent complications associated with the use of the vaccine.

Immunofluorescence detection of the presence of S-protein in the tissues and organs of mice showed its presence in mice of the experimental group (see Fig. 10B - skeletal muscles of mouse 3.1, withdrawn from the experiment on the 4th week after the administration of the vaccine), as well as in the tissue of the positive control (see Fig. 10A - lung tissue of a patient who died from a verified SARS-CoV-2 infection), and absence in the negative control group (see Fig. 10B - skeletal muscles of mouse 5.6, which was not administered the vaccine), which is direct evidence of the synthesis of S-protein as a result of the administration of the vector vaccine encoding the S-protein of the SARS-CoV-2 virus, according to the invention.

Example 7. Study of the immunogenicity of experimental adeno-associated vectors of Spike protein modification variants 2, 3 and 4 in vivo

The study was conducted on BALB/c mice, females, 6-8 weeks old. The mice were obtained from the Stolbovaya nursery of the Federal State Budgetary Scientific Institution "National Center for Biomedical Technologies" of the Federal Medical and Biological Agency of Russia, Moscow Region. All experiments involving laboratory animals were approved by the Bioethics Committee of the A.A. Smorodintsev Research Institute of Influenza of the Ministry of Health of the Russian Federation, Protocol No. 02 dated January 24, 2023.

Animals (50 animals in total) were immunized after the end of quarantine by intramuscular injection of 100 μl of the preparation containing 10 ¹⁰ or 10 ¹¹ GC of adeno-associated vectors in accordance with the study design into the hind paw. All animals were divided into 7 groups (see Table 5 for details):

1-2 - immunization with the rAAV9-SARS2 vaccine (Spike) at a dose of 10 ¹⁰ or 10 ¹¹ GC,

3-4 - immunization with rAAV9-SARS3 (S1) vaccine at a dose of 10 ¹⁰ or 10 ¹¹ GC,

5-6 - immunization with rAAV9-SARS4 vaccine (RBD) at a dose of 10 ¹⁰ or 10 ¹¹ GC,

7 - immunization was not carried out (intact).

The body weight and condition of the animals were monitored for 2 weeks, weighing them once every 2-3 days. After 4 and 12 weeks, all animals had their blood serum taken during their lifetime to detect virus-specific IgG antibodies and neutralizing antibodies.

Table 5. Design of the study on laboratory mice

№ gr Preparation Dose Number of mice Observations and manipulations by day 0 1-2 weeks Week 4 Week 12 1 Spike 10 ¹¹ GK 7 B, I (7) B(7) C(7) C(7), T(7) 2 Spike 10 ¹⁰ GK 7 B, I (7) B(7) C(7) C(7), T(7) 3 S1 10 ¹¹ GK 7 B, I (7) B(7) C(7) C(7), T(7) 4 S1 10 ¹⁰ GK 7 B, I (7) B(7) C(7) C(7), T(7) 5 RBD 10 ¹¹ GK 7 B, I (7) B(7) C(7) C(7), T(7) 6 RBD 10 ¹⁰ GK 7 B, I (7) B(7) C(7) C(7), T(7) 7 Intact - 8 In (8) In (8) C (8) C(8), T(8)

Note: GC - genomic copies, and - immunization, B - weighing (once every 2-3 days), C - serum sampling, O - collection of organs and tissues for IHC, T - Spleen collection for T-cell response. The number of animals is given in brackets.

During the 2 weeks after immunization, the weight of mice fluctuated within ±5% of the initial weight. The exception was a short-term drop in weight in 3/7 mice in group 6 (RBD, dose 10 ¹⁰ ), observed on days 13, 14 (see Fig. 11).

The presence of virus-neutralizing antibodies was determined in animal sera using the classical RMN method [15], [16] using the SARS-CoV-2 Delta virus (hCoV-19/Russia/SPE-RII-27029V3/2021 strain, line B.1.617.3, GISAID database number: EPI_ISL_8112507) at a dose of 100 TCID50 per well. The neutralizing titer was considered to be the highest serum dilution at which there were no signs of CPE in the well. The study of the antibody response in vaccinated animals showed that virus-neutralizing antibodies are formed only in mice vaccinated with the rAAV9-SARS2 (Spike) construct at both doses and in some mice (2/7) vaccinated with the rAAV9-SARS4 (RBD) construct at a high dose. In mice vaccinated with the rAAV9-SARS3 (S1) construct, virus-neutralizing antibodies are not formed. However, antibody titers in mice vaccinated with the rAAV9-SARS2 (Spike) construct statistically significantly increased by the 12th week after immunization and had a direct dose dependence (see Fig. 12 and Table 6).

Table 6. Geometric mean values of antibody titers in groups of immunized animals

Group Neutralization IFA_SARS-CoV-2 IFA_AAV9 4 weeks 12 weeks 4 weeks 12 weeks 4 weeks 12 weeks 1 Spike 10 ¹¹ 97.5* 640.0* 3901 28265 689* 312** 2 Spike 10 ¹⁰ 5.0* 59.4* 181 4755 95* 78** 3 S1 10 ¹¹ 5.0* 5.0* 221 362 689* 172* 4 S1 10 ¹⁰ 5.0 5.0 100 110 14* 13* 5 RBD 10 ¹¹ 6.7* 6.7* 221 328 464* 128* 6 RBD 10 ¹⁰ 5.0 5.0 100 100 20* 32* 7 Intact 5.0 5.0 100 100 8 8

Note: Asterisks indicate selected statistically significant differences when comparing doses: *p<0.0001; **p=0.0271;

The presence of virus-specific IgG antibodies was determined by ELISA with samples of whole animal serum. The antigen used was a preparation of the inactivated sucrose gradient-purified SARS-CoV-2 Beta virus or purified capsids of the adeno-associated vector type 9 (AAV9 empty capsid, Progen).

Inactivated coronavirus was adsorbed at a concentration of 5 μg/ml, purified AAV9 at a concentration of 3*10 ⁸ capsids/ml in PBS (Biolot, Russia) overnight at +4°C in 96-well Maxosorp plates (Nunc) (100 μl/well).

In contrast to neutralizing antibodies, virus-specific IgG antibodies were observed in animals vaccinated with all three constructs, but for rAAV9-SARS3 (S1) and rAAV9-SARS4 (RBD) vectors only at a high dose (see Fig. 13 and Table 6). Antibody titers were directly dependent on the insert length: the highest antibody level was observed in mice vaccinated with rAAV9-SARS2 (Spike), the lowest - in mice vaccinated with rAAV9-SARS4 (RBD). Antibody titers increased in the period of 4-12 weeks, the difference was statistically significant for the rAAV9-SARS2 (Spike) construct.

The humoral response to AAV9 capsid antigen was dose-dependent at 4 weeks post-vaccination, with no differences between the different vector groups (see Fig. 14 and Table 6). At 12 weeks post-immunization, statistically significant decreases in AAV9 antibodies were observed in the groups of mice inoculated with the high dose rAAV9-SARS3 (S1) and rAAV9-SARS4 (RBD) constructs. A decrease in antibody titers was also observed in the group of mice inoculated with the high dose rAAV9-SARS2 (Spike) construct, but this decrease was not statistically significant. In the remaining groups, AAV9 antibody titers remained virtually unchanged over the period 4-12 weeks.

Example 8. Study of the T-cell immune response of experimental proteins in the context of an AAV9-based vaccine during immunization of mice in vivo

The study was performed on groups of animals as described in Example 7. The systemic antigen-specific T-cell response was determined 12 weeks after vaccination. Spleens were collected from all animals, followed by isolation of splenocytes.

Specific stimulation was performed with PepTivator S (Miltenyi Biotec, a cocktail of overlapping peptides consisting of 11 peptides covering the complete sequence of the SARS - CoV-2 membrane glycoprotein (GenBank MN908947.3)) for 24 h at 37°C and 5% _CO2 in the presence of the cellular transport inhibitor brefeldin A (GolgiPlug included in the CytoFix/CytoPerm intracellular staining kit (BD)) and co-stimulating antibodies to CD28 (Biolegend).

Complete culture medium containing brefeldin A and costimulatory antibodies to CD28 was used as an unstimulated control, and a mixture of PMA (Sigma) and ionomycin (Sigma) was used as a positive stimulation control.

Response was assessed by the relative numbers of IFNγ/IL2/TNFα cytokine-producing CD4+ and CD8+ effector memory T cells measured by flow cytometry after in vitro stimulation of splenocytes with a cocktail of overlapping S protein peptides (PepTivator S).

Immunization with the studied drugs induced predominantly a CD8+ effector T cell response, the intensity of which was determined by the size of the insert expressed by the adeno-associated vector (Fig. 15A, B).

In this case, the main proportion of antigen-specific effector CD8+ T-lymphocytes was made up of the population of dual (IFNγ+IL-2-TNFα+) producers (Fig. 16B, G, E). The CD4+ antigen-specific T-cell response was observed only in the group immunized with the adeno-associated vector rAAV9-SARS2 expressing the full-length Spike protein, and was characterized by the predominance of monofunctional IFNγ+IL-2-TNFα- Tem cells (see Fig. 16A, B, D).

Analysis of total cytokine production (IFNγ/IL2/TNFα) did not reveal a pronounced dose-dependent effect for all the studied constructs. Differences were noted when analyzing the proportion of IFNγ+IL-2-TNFα+ CD8+ Tem lymphocytes. In the groups immunized with the adeno-associated vector rAAV9-SARS2 expressing the full-length Spike protein, a direct relationship was observed between the dose and the relative number of IFNγ+IL-2-TNFα+ CD8+ Tem cells, and an inverse relationship was observed in the groups immunized with the rAAV9-SARS3 (S1) and rAAV9-SARS4 (RBD) constructs.

Thus, based on the results of the study, it was found that vector vaccine preparations are characterized by different levels of immunogenicity depending on the design and the administered dose. A higher dose of the preparation and a longer antigen insert were associated with a higher level of both humoral and T-cell responses to the S protein of the SARS-CoV-2 virus.

Virus-neutralizing antibodies were detected only in animals vaccinated with the construct carrying the full-length rAAV9-SARS2 spike protein (Spike), the antibody titer increased in the period 4-12 weeks after vaccination and had a direct dose-dependent effect. Virus-specific IgG antibodies were observed in animals vaccinated with all three variants of the constructs, but for vectors carrying truncated versions of the S protein (rAAV9-SARS3 (S1) and rAAV9-SARS4 (RBD)) - only at a high dose. IgG antibody titers had a direct dependence on the insert length and increased in the period 4-12 weeks. The level of IgG antibodies to the vector (AAV9 capsid) 4 and 12 weeks after immunization directly depended on the administered dose and did not depend on the construct.

It was also found that immunization of animals with the developed vector vaccine preparations leads to the formation of a pronounced CD8+ T-cell response to the peptides of the S-protein of the SARS-CoV-2 virus, the intensity of which does not have a clear dose dependence, but directly depends on the size of the insert. In the population of antigen-specific CD8+ effector memory cells, double positive cells (IFNg+TNFa+) predominated. An antigen-specific CD4+ T-cell response was recorded only in mice vaccinated with a construct carrying the full-length rAAV9-SARS2 spike protein (Spike), while IFNg-producing cells predominated in the population.

The experiments show that although the use of the S1 subunit and the RBD domain have proven effective in previously developed vaccines against the SARS-CoV-2 virus, these constructs have proven insufficiently effective in the vector vaccines being developed and cannot be used for effective prevention of coronavirus infection caused by the SARS-CoV-2 virus. At the same time, a vaccine based on the adeno-associated virus of serotype 9 encoding a full-length S protein is capable of inducing activation of not only the humoral, but also the cellular immune response after a single intramuscular administration, while after one injection, the expression of the construct continues for more than 24 weeks. The safety of the developed vector vaccine is ensured by local production of the antigen at the injection site.

Thus, as a result of the conducted research, a vaccine based on the adeno-associated virus 9 serotype was developed, providing effective stimulation of the humoral and cellular response, which can be used to prevent coronavirus infection caused by the SARS-COV-2 virus.

Although the invention has been described with reference to the disclosed embodiments, it should be obvious to those skilled in the art that the specific experiments described in detail are provided merely for the purpose of illustrating the present invention and should not be considered as limiting the scope of the invention in any way. It should be understood that various modifications can be made without departing from the spirit of the present invention.

Cited documents:

[1] Travieso T. et al., “The use of viral vectors in vaccine development,” pp. 1 - 10, 2022, doi: 10.1038/s41541-022-00503-y.

[2] Qin X. et al., “Development of an SARS-CoV-2 Vaccine and Its Immunogenicity in Mice,” vol. 12, no. March, pp. 1 - 10, 2022, doi: 10.3389/fcimb.2022.802147.

[3] Wu F. et al., “Since January 2020 Elsevier has created a COVID-19 resource center with free information in English and Mandarin on the novel coronavirus COVID- research that is available on the COVID-19 resource center - including this Single-shot AAV-vectored vaccine against SARS-CoV-2 with fast and long-lasting immunity,” no. January, 2020.

[4] Maciorowski D. et al., “Immunogenicity of an AAV-Based COVID-19 Vaccine in Murine Models of Obesity and Aging,” pp. 1 - 12, 2022.

[5] Zabaleta N. et al., “Durable immunogenicity, adaptation to emerging variants, and low-dose efficacy of an AAV-based COVID-19 vaccine platform in macaques,” vol. 30, no. 9, pp. 2952 - 2967, 2022, doi: 10.1016/j.ymthe.2022.05.007.

[6] NCT05037188, “Clinical Study of the Safety and Immunogenicity of a Recombinant Viral Vector AAV5 (Adeno-Associated Virus Type 5)-RBD (Receptor Binding Domain)-S Vaccine for the Prevention of Coronavirus Infection (COVID-19) (COVER).” [Online]. Available: https://clinicaltrials.gov/ct2/show/NCT05037188?term=Adeno-Associated&cond=COVID-19&draw=2&rank=1

[7] Xia S. et al., “The role of furin cleavage site in SARS-CoV-2 spike protein-mediated membrane fusion in the presence or absence of trypsin,” Sig Transduct Target Ther 5, 92 (2020). https://doi.org/10.1038/s41392-020-0184-0

[8] Zhang Y. et al., “A second functional furin site in the SARS-CoV-2 spike protein,” Emerg Microbes Infect. 2022; 11(1): 182 - 194, Published online 2022 Jan 4. doi: 10.1080/22221751.2021.2014284

[9] Pedenko B. et al., “SARS-CoV-2 S Glycoprotein Stabilization Strategies”, Viruses 2023, 15(2), 558. https://doi.org/10.3390/v15020558

[10] Zhang J. et al., “Structural impact on SARS-CoV-2 spike protein by D614G substitution”, Science, 16 Mar 2021, 372(6541): pp.525-530, doi: 10.1126/science.abf2303

[11] Zincarelli C. et al. “Analysis of AAV serotypes 1-9 mediated gene expression and tropism in mice after systemic injection”, Mol Ther. 2008 Jun; 16(6):1073-80. doi: 10.1038/mt.2008.76. Epub 2008 Apr 15.

[12] Rajendra Y. et al., “Transcriptional and post-transcriptional limitations of high-yielding, PEI-mediated transient transfection with CHO and HEK-293E cells.” Biotechnol Prog. 2015;31(2):541 - 9. https://doi.org/10.1002/btpr.2064

[13] Joshi P.R.H. et al., “Advances in molecular design and bioprocessing of recombinant adeno-associated virus gene delivery vectors using the insect-cell baculovirus expression platform,” Biotechnol. J. 2020;16

[14] Robert M.A. et al., “Manufacturing of recombinant adeno-associated viruses using mammalian expression platforms”, Biotechnol. J. 2017;12. doi: 10.1002/biot.201600193.

[15] Amanat F. et al., “An In Vitro Microneutralization Assay for SARS-CoV-2 Serology and Drug Screening,” Curr. Protoc. Microbiol., vol. 58, no. 1, pp. 1 - 15, 2020, doi: 10.1002/cpmc.108.

[16] Manenti A. et al., “Evaluation of SARS-CoV-2 neutralizing antibodies using a CPE-based colorimetric live virus micro-neutralization assay in human serum samples,” J. Med. Virol., vol. 92, no. 10, pp. 2096 - 2104, 2020, doi: 10.1002/jmv.25986.

This XML file does not appear to have any style information associated with it. The document tree is shown below.

--->

<ST26SequenceListing dtdVersion="V1_3" fileName="11.10.2024_List

sequences.xml" softwareName="WIPO Sequence"

softwareVersion="2.3.0" productionDate="2024-10-11">

<ApplicantFileReference>4117646</ApplicantFileReference>

<ApplicantName languageCode="en">Limited liability company

responsibility of Swiftgen</ApplicantName>

<ApplicantNameLatin>Swiftgen LLC</ApplicantNameLatin>

<InventionTitle languageCode="en">Vaccine based on

adeno-associated vector type 9 encoding Spike protein, for

prevention of coronavirus infection SARS-CoV-2</InventionTitle>

<SequenceTotalQuantity>27</SequenceTotalQuantity>

<SequenceData sequenceIDNumber="1">

<INSDSeq>

<INSDSeq_length>1273</INSDSeq_length>

<INSDSeq_moltype>AA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..1273</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>protein</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q2">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>Betacoronavirus</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>MFVFLVLLPLVSSQCVNLTTRTQLPPAYTNSFTRGVYYPDKVFRSSVLHSTQ

DLFLPFFSNVTWFHAIHVSGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNN

ATNVVIKVCEFQFCNDPFLGVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREF

VFKNIDGYFKIYSKHTPINLVRDLPQGFSALEPLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGA

AAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNI

TNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVI

RGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEI

YQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFN

FNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLY

QDVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEHVNNSYECDIPIGAGICASYQTQTNSPR

RARSVASQSIIAYTMSLGAENSVAYSNNSIAIPTNFTISVTTEILPVSMTKTSVDCTMYICGDSTECSNL

LLQYGSFCTQLNRALTGIAVEQDKNTQEVFAQVKQIYKTPPIKDFGGFNFSQILPDPSKPSKRSFIEDLL

FNKVTLADAGFIKQYGDCLGDIAARDLICAQKFNGLTVLPPLLTDEMIAQYTSALLAGTITSGWTFGAGA

ALQIPFAMQMAYRFNGIGVTQNVLYENQKLIANQFNSAIGKIQDSLSSTASALGKLQDVVNQNAQALNTL

VKQLSSNFGAISSVLNDILSRLDKVEAEVQIDRLITGRLQSLQTYVTQQLIRAAEIRASANLAATKMSEC

VLGQSKRVDFCGKGYHLMSFPQSAPHGVVFLHVTYVPAQEKNFTTAPAICHDGKAHFPREGVFVSNGTHW

FVTQRNFYEPQIITTDNTFVSGNCDVVIGIVNNTVYDPLQPELDSFKEELDKYFKNHTSPDVDLGDISGI

NASVVNIQKEIDRLNEVAKNLNESLIDLQELGKYEQYIKWPWYIWLGFIAGLIAIVMVTIMLCCMTSCCS

CLKGCCSCGSCCKFDEDDSEPVLKGVKLHYT</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="2">

<INSDSeq>

<INSDSeq_length>1270</INSDSeq_length>

<INSDSeq_moltype>AA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..1270</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>protein</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q4">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>MFVFLVLLPLVSSQCVNLTTRTQLPPAYTNSFTRGVYYPDKVFRSSVLHSTQ

DLFLPFFSNVTWFHAIHVSGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNN

ATNVVIKVCEFQFCNDPFLGVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREF

VFKNIDGYFKIYSKHTPINLVRDLPQGFSALEPLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGA

AAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNI

TNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVI

RGDEVRQIAPGQTGKIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEI

YQAGSTPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFN

FNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLY

QGVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEHVNNSYECDIPIGAGICASYQTQTNSPA

SVASQSIIAYTMSLGAENSVAYSNNSIAIPTNFTISVTTEILPVSMTKTSVDCTMYICGDSTECSNLLLQ

YGSFCTQLNRALTGIAVEQDKNTQEVFAQVKQIYKTPPIKDFGGFNFSQILPDPSKPSKRSFIEDLLFNK

VTLADAGFIKQYGDCLGDIAARDLICAQKFNGLTVLPPLLTDEMIAQYTSALLAGTITSGWTFGAGAALQ

IPFAMQMAYRFNGIGVTQNVLYENQKLIANQFNSAIGKIQDSLSSTASALGKLQDVVNQNAQALNTLVKQ

LSSNFGAISSVLNDILSRLDPPEAEVQIDRLITGRLQSLQTYVTQQLIRAAEIRASANLAATKMSECVLG

QSKRVDFCGKGYHLMSFPQSAPHGVVFLHVTYVPAQEKNFTTAPAICHDGKAHFPREGVFVSNGTHWFVT

QRNFYEPQIITTDNTFVSGNCDVVIGIVNNTVYDPLQPELDSFKEELDKYFKNHTSPDVDLGDISGINAS

VVNIQKEIDRLNEVAKNLNESLIDLQELGKYEQYIKWPWYIWLGFIAGLIAIVMVTIMLCCMTSCCSCLK

GCCSCGSCCKFDEDDSEPVLKGVKLHYT</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="3">

<INSDSeq>

<INSDSeq_length>1270</INSDSeq_length>

<INSDSeq_moltype>AA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..1270</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>protein</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q6">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>MFVFLVLLPLVSSQCVNLTTRTQLPPAYTNSFTRGVYYPDKVFRSSVLHSTQ

DLFLPFFSNVTWFHAIHVSGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNN

ATNVVIKVCEFQFCNDPFLGVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREF

VFKNIDGYFKIYSKHTPINLVRDLPQGFSALEPLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGA

AAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNI

TNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVI

RGDEVRQIAPGQTGNIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEI

YQAGSTPCNGVKGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFN

FNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLY

QGVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEHVNNSYECDIPIGAGICASYQTQTNSPA

SVASQSIIAYTMSLGAENSVAYSNNSIAIPTNFTISVTTEILPVSMTKTSVDCTMYICGDSTECSNLLLQ

YGSFCTQLNRALTGIAVEQDKNTQEVFAQVKQIYKTPPIKDFGGFNFSQILPDPSKPSKRSFIEDLLFNK

VTLADAGFIKQYGDCLGDIAARDLICAQKFNGLTVLPPLLTDEMIAQYTSALLAGTITSGWTFGAGAALQ

IPFAMQMAYRFNGIGVTQNVLYENQKLIANQFNSAIGKIQDSLSSTASALGKLQDVVNQNAQALNTLVKQ

LSSNFGAISSVLNDILSRLDPPEAEVQIDRLITGRLQSLQTYVTQQLIRAAEIRASANLAATKMSECVLG

QSKRVDFCGKGYHLMSFPQSAPHGVVFLHVTYVPAQEKNFTTAPAICHDGKAHFPREGVFVSNGTHWFVT

QRNFYEPQIITTDNTFVSGNCDVVIGIVNNTVYDPLQPELDSFKEELDKYFKNHTSPDVDLGDISGINAS

VVNIQKEIDRLNEVAKNLNESLIDLQELGKYEQYIKWPWYIWLGFIAGLIAIVMVTIMLCCMTSCCSCLK

GCCSCGSCCKFDEDDSEPVLKGVKLHYT</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="4">

<INSDSeq>

<INSDSeq_length>677</INSDSeq_length>

<INSDSeq_moltype>AA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..677</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>protein</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q8">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>MFVFLVLLPLVSSQCVNLTTRTQLPPAYTNSFTRGVYYPDKVFRSSVLHSTQ

DLFLPFFSNVTWFHAIHVSGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNN

ATNVVIKVCEFQFCNDPFLGVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREF

VFKNIDGYFKIYSKHTPINLVRDLPQGFSALEPLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGA

AAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNI

TNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVI

RGDEVRQIAPGQTGNIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEI

YQAGSTPCNGVKGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFN

FNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLY

QGVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEHVNNSYECDIPIGAGICASYQTQ</INS

DSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="5">

<INSDSeq>

<INSDSeq_length>393</INSDSeq_length>

<INSDSeq_moltype>AA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..393</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>protein</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q10">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>MFVFLVLLPLVSSQTLKSFTVEKGIYQTSNFRVQPTESIVRFPNITNLCPFG

EVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQ

IAPGQTGNIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGSTP

CNGVKGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFNFNGLTGT

GVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLYQGVNCTE

VPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEHVNNSYECDIPIGAGICASYQTQTNS</INSDSeq

_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="6">

<INSDSeq>

<INSDSeq_length>1270</INSDSeq_length>

<INSDSeq_moltype>AA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..1270</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>protein</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q12">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>MFVFLVLLPLVSSQCVNLTTRTQLPPAYTNSFTRGVYYPDKVFRSSVLHSTQ

DLFLPFFSNVTWFHAIHVSGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIVNN

ATNVVIKVCEFQFCNDPFLGVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMDLEGKQGNFKNLREF

VFKNIDGYFKIYSKHTPINLVRDLPQGFSALEPLVDLPIGINITRFQTLLALHRSYLTPGDSSSGWTAGA

AAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSETKCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNI

TNLCPFGEVFNATRFASVYAWNRKRISNCVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVI

RGDEVRQIAPGQTGNIADYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYRYRLFRKSNLKPFERDISTEI

YQAGSKPCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKNKCVNFN

FNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLY

QGVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEHVNNSYECDIPIGAGICASYQTQTNSPA

SVASQSIIAYTMSLGAENSVAYSNNSIAIPTNFTISVTTEILPVSMTKTSVDCTMYICGDSTECSNLLLQ

YGSFCTQLNRALTGIAVEQDKNTQEVFAQVKQIYKTPPIKDFGGFNFSQILPDPSKPSKRSFIEDLLFNK

VTLADAGFIKQYGDCLGDIAARDLICAQKFNGLTVLPPLLTDEMIAQYTSALLAGTITSGWTFGAGAALQ

IPFAMQMAYRFNGIGVTQNVLYENQKLIANQFNSAIGKIQDSLSSTASALGKLQDVVNQNAQALNTLVKQ

LSSNFGAISSVLNDILSRLDPPEAEVQIDRLITGRLQSLQTYVTQQLIRAAEIRASANLAATKMSECVLG

QSKRVDFCGKGYHLMSFPQSAPHGVVFLHVTYVPAQEKNFTTAPAICHDGKAHFPREGVFVSNGTHWFVT

QRNFYEPQIITTDNTFVSGNCDVVIGIVNNTVYDPLQPELDSFKEELDKYFKNHTSPDVDLGDISGINAS

VVNIQKEIDRLNEVAKNLNESLIDLQELGKYEQYIKWPWYIWLGFIAGLIAIVMVTIMLCCMTSCCSCLK

GCCSCGSCCKFDEDDSEPVLKGVKLHYT</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="7">

<INSDSeq>

<INSDSeq_length>3813</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..3813</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q14">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>atgttcgtgtttctggtgctgctgcctctggtgtccagccagtgtgtgaacc

tgaccacaagaacccagctgcctccagcctacaccaacagctttaccagaggcgtgtactaccccgacaa

ggtgttcagatccagcgtgctgcactctacccaggacctgttcctgcctttcttcagcaacgtgacctgg

ttccacgccatccacgtgtccggcaccaatggcaccaagagattcgacaaccccgtgctgcccttcaacg

acggggtgtactttgccagcaccgagaagtccaacatcatcagaggctggatcttcggcaccacactgga

cagcaagacccagcctgctgatcgtgaacaacgccaccaacgtggtcatcaaagtgtgcgagttccag

ttctgcaacgaccccttcctgggcgtctactatcacaagaacaacaagagctggatggaaagcgagttcc

gggtgtacagcagcgccaacaactgcaccttcgagtacgtgtcccagcctttcctgatggacctggaagg

caagcagggcaacttcaagaacctgcgcgagttcgtgttcaagaacatcgacggctacttcaagatctac

agcaagcacacccctatcaacctcgtgcgggatctgcctcagggcttctctgctctggaacccctggtgg

atctgcccatcggcatcaacatcacccggtttcagacactgctggccctgcacagaagctacctgacacc

tggcgatagcagcagcggatggacagctggtgccgccgcttactatgtgggctacctgcagcctagaacc

ttcctgctgaagtacaacgagaacggcaccatcaccgacgccgtggattgtgctctggatcctctgagcg

agacaaagtgcaccctgaagtccttcaccgtggaaaagggcatctaccagaccagcaacttccgggtgca

gcccaccgaatccatcgtgcggttccccaatatcaccaatctgtgccccttcggcgaggtgttcaatgcc

accagatcgcctctgtgtacgcctggaaccggaagcggatcagcaattgcgtggccgactactccgtgc

tgtacaactccgccagcttcagcaccttcaagtgctacggcgtgtcccctaccaagctgaacgacctgtg

cttcacaaacgtgtacgccgacagcttcgtgatccggggagatgaagtgcggcagattgcccctggacag

acaggcaagatcgccgactacaactacaagctgcccgacgacttcaccggctgtgtgattgcctggaaca

gcaacaacctggactccaaagtcggcggcaactacaattacctgtaccggctgttccggaagtccaatct

gaagcccttcgagcgggacatctccaccgagatctatcaggccggcagcaccccttgtaacggcgtggaa

ggcttcaactgctacttcccactgcagtcctacggctttcagcccacaaatggcgtgggctatcagccct

acagagtggtggtgctgagcttcgaactgctgcatgcccctgccacagtgtgcggccctaagaaaagcac

caatctcgtgaagaacaaatgcgtgaacttcaacttcaacggcctgaccggcaccggcgtgctgacagag

agcaacaagaagttcctgccattccagcagtttggccgggatatcgccgataccacagacgccgttagag

atccccagacactggaaatcctggacatcaccccttgcagcttcggcggagtgtctgtgatcacccctgg

caccaacaccagcaatcaggtggcagtgctgtaccagggcgtgaactgtaccgaagtgcccgtggccatt

cacgccgatcagctgacacctacatggcgggtgtactccaccggcagcaatgtgtttcagaccagagccg

gctgtctgatcggagccgagcacgtgaacaatagctacgagtgcgacatccccatcggcgctggcatctg

tgccagctaccagacacagacaaacagccctgcttctgtggccagccagagcatcattgcctacacaatg

tctctgggcgccgagaacagcgtggcctactccaacaactctatcgctatccccaccaacttcaccatca

gcgtgaccacagatcctgcctgtgtccatgaccaagaccagcgtggactgcaccatgtacatctgcgg

cgattccaccgagtgctccaacctgctgctgcagtacggcagcttctgcacccagctgaatagagccctg

acagggatcgccgtggaacaggacaagaacacccaagaggtgttcgcccaagtgaagcagatctacaaga

cccctcctatcaaggacttcggcggcttcaatttcagccagattctgcccgatcctagcaagcccagcaa

gcggagcttcatcgaggacctgctgttcaacaaagtgacactggccgacgccggcttcatcaagcagtat

ggcgattgtctgggcgacattgccgccagggatctgatttgcgcccagaagtttaacggactgacagtgc

tgccaccactgctgaccgatgagatgatcgcccagtacacatctgccctgctggccggcacaatcacaag

cggctggacatttggagctggcgccgctctgcagatcccctttgctatgcagatggcctaccggttcaac

ggcatcggagtgacccagaatgtgctgtacgagaaccagaagctgatcgccaaccagttcaacagcgcca

tcggcaagatccaggacagcctgagcagcacagcaagcgccctgggaaagctgcaggacgtggtcaacca

gaatgcccaggcactgaacaccctggtcaagcagctgtcctccaacttcggcgccatcagctctgtgctg

aacgatatcctgagcagactggaccctccggaagccgaggtgcagatcgacagactgatcaccggaaggc

tgcagtccctgcagacctacgttacccagcagctgatcagagccgccgagattagagcctctgccaatct

ggccgccaccaagatgtctgagtgtgtgctgggccagagcaagagagtggacttttgcggcaagggctac

cacctgatgagcttccctcagtctgcccctcacggcgtggtgtttctgcacgtgacatacgtgcccgctc

aagagaagaatttcaccaccgctccagccatctgccacgacggcaaagcccactttcctagagaaggcgt

gttcgtgtccaacggcacccattggttcgtgacccagcggaacttctacgagccccagatcatcaccacc

gacaacaccttcgtgtctggcaactgcgacgtcgtgatcggcattgtgaacaataccgtgtacgaccctc

tgcagcccgagctggacagcttcaaagaggaactggataagtactttaagaaccacacaagccccgacgt

ggacctgggcgatatcagcggaatcaatgccagcgtcgtgaacatccagaaagagatcgaccggctgaac

gaggtggccaagaatctgaacgagagcctgatcgacctgcaagaactggggaagtacgagcagtacatca

agtggccctggtacatctggctggggctttatcgccggactgattgccatcgtgatggtcacaatcatgct

gtgttgcatgaccagctgctgtagctgcctgaagggctgttgtagctgtggcagctgctgcaagttcgac

gaggacgattctgagcccgtgctgaagggcgtgaaactgcactacacatga</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="8">

<INSDSeq>

<INSDSeq_length>3813</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..3813</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q16">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>atgttcgtgtttctggtgctgctgcctctggtgtccagccagtgtgtgaacc

tgaccacaagaacccagctgcctccagcctacaccaacagctttaccagaggcgtgtactaccccgacaa

ggtgttcagatccagcgtgctgcactctacccaggacctgttcctgcctttcttcagcaacgtgacctgg

ttccacgccatccacgtgtccggcaccaatggcaccaagagattcgacaaccccgtgctgcccttcaacg

acggggtgtactttgccagcaccgagaagtccaacatcatcagaggctggatcttcggcaccacactgga

cagcaagacccagcctgctgatcgtgaacaacgccaccaacgtggtcatcaaagtgtgcgagttccag

ttctgcaacgaccccttcctgggcgtctactatcacaagaacaacaagagctggatggaaagcgagttcc

gggtgtacagcagcgccaacaactgcaccttcgagtacgtgtcccagcctttcctgatggacctggaagg

caagcagggcaacttcaagaacctgcgcgagttcgtgttcaagaacatcgacggctacttcaagatctac

agcaagcacacccctatcaacctcgtgcgggatctgcctcagggcttctctgctctggaacccctggtgg

atctgcccatcggcatcaacatcacccggtttcagacactgctggccctgcacagaagctacctgacacc

tggcgatagcagcagcggatggacagctggtgccgccgcttactatgtgggctacctgcagcctagaacc

ttcctgctgaagtacaacgagaacggcaccatcaccgacgccgtggattgtgctctggatcctctgagcg

agacaaagtgcaccctgaagtccttcaccgtggaaaagggcatctaccagaccagcaacttccgggtgca

gcccaccgaatccatcgtgcggttccccaatatcaccaatctgtgccccttcggcgaggtgttcaatgcc

accagatcgcctctgtgtacgcctggaaccggaagcggatcagcaattgcgtggccgactactccgtgc

tgtacaactccgccagcttcagcaccttcaagtgctacggcgtgtcccctaccaagctgaacgacctgtg

cttcacaaacgtgtacgccgacagcttcgtgatccggggagatgaagtgcggcagattgcccctggacag

acaggcaacatcgccgactacaactacaagctgcccgacgacttcaccggctgtgtgattgcctggaaca

gcaacaacctggactccaaagtcggcggcaactacaattacctgtaccggctgttccggaagtccaatct

gaagcccttcgagcgggacatctccaccgagatctatcaggccggcagcaccccttgtaacggcgtgaaa

ggcttcaactgctacttcccactgcagtcctacggctttcagcccacaaatggcgtgggctatcagccct

acagagtggtggtgctgagcttcgaactgctgcatgcccctgccacagtgtgcggccctaagaaaagcac

caatctcgtgaagaacaaatgcgtgaacttcaacttcaacggcctgaccggcaccggcgtgctgacagag

agcaacaagaagttcctgccattccagcagtttggccgggatatcgccgataccacagacgccgttagag

atccccagacactggaaatcctggacatcaccccttgcagcttcggcggagtgtctgtgatcacccctgg

caccaacaccagcaatcaggtggcagtgctgtaccagggcgtgaactgtaccgaagtgcccgtggccatt

cacgccgatcagctgacacctacatggcgggtgtactccaccggcagcaatgtgtttcagaccagagccg

gctgtctgatcggagccgagcacgtgaacaatagctacgagtgcgacatccccatcggcgctggcatctg

tgccagctaccagacacagacaaacagccctgcttctgtggccagccagagcatcattgcctacacaatg

tctctgggcgccgagaacagcgtggcctactccaacaactctatcgctatccccaccaacttcaccatca

gcgtgaccacagatcctgcctgtgtccatgaccaagaccagcgtggactgcaccatgtacatctgcgg

cgattccaccgagtgctccaacctgctgctgcagtacggcagcttctgcacccagctgaatagagccctg

acagggatcgccgtggaacaggacaagaacacccaagaggtgttcgcccaagtgaagcagatctacaaga

cccctcctatcaaggacttcggcggcttcaatttcagccagattctgcccgatcctagcaagcccagcaa

gcggagcttcatcgaggacctgctgttcaacaaagtgacactggccgacgccggcttcatcaagcagtat

ggcgattgtctgggcgacattgccgccagggatctgatttgcgcccagaagtttaacggactgacagtgc

tgccaccactgctgaccgatgagatgatcgcccagtacacatctgccctgctggccggcacaatcacaag

cggctggacatttggagctggcgccgctctgcagatcccctttgctatgcagatggcctaccggttcaac

ggcatcggagtgacccagaatgtgctgtacgagaaccagaagctgatcgccaaccagttcaacagcgcca

tcggcaagatccaggacagcctgagcagcacagcaagcgccctgggaaagctgcaggacgtggtcaacca

gaatgcccaggcactgaacaccctggtcaagcagctgtcctccaacttcggcgccatcagctctgtgctg

aacgatatcctgagcagactggaccctccggaagccgaggtgcagatcgacagactgatcaccggaaggc

tgcagtccctgcagacctacgttacccagcagctgatcagagccgccgagattagagcctctgccaatct

ggccgccaccaagatgtctgagtgtgtgctgggccagagcaagagagtggacttttgcggcaagggctac

cacctgatgagcttccctcagtctgcccctcacggcgtggtgtttctgcacgtgacatacgtgcccgctc

aagagaagaatttcaccaccgctccagccatctgccacgacggcaaagcccactttcctagagaaggcgt

gttcgtgtccaacggcacccattggttcgtgacccagcggaacttctacgagccccagatcatcaccacc

gacaacaccttcgtgtctggcaactgcgacgtcgtgatcggcattgtgaacaataccgtgtacgaccctc

tgcagcccgagctggacagcttcaaagaggaactggataagtactttaagaaccacacaagccccgacgt

ggacctgggcgatatcagcggaatcaatgccagcgtcgtgaacatccagaaagagatcgaccggctgaac

gaggtggccaagaatctgaacgagagcctgatcgacctgcaagaactggggaagtacgagcagtacatca

agtggccctggtacatctggctggggctttatcgccggactgattgccatcgtgatggtcacaatcatgct

gtgttgcatgaccagctgctgtagctgcctgaagggctgttgtagctgtggcagctgctgcaagttcgac

gaggacgattctgagcccgtgctgaagggcgtgaaactgcactacacatga</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="9">

<INSDSeq>

<INSDSeq_length>2034</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..2034</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q18">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>atgttcgtgtttctggtgctgctgcctctggtgtccagccagtgtgtgaacc

tgaccacaagaacccagctgcctccagcctacaccaacagctttaccagaggcgtgtactaccccgacaa

ggtgttcagatccagcgtgctgcactctacccaggacctgttcctgcctttcttcagcaacgtgacctgg

ttccacgccatccacgtgtccggcaccaatggcaccaagagattcgacaaccccgtgctgcccttcaacg

acggggtgtactttgccagcaccgagaagtccaacatcatcagaggctggatcttcggcaccacactgga

cagcaagacccagcctgctgatcgtgaacaacgccaccaacgtggtcatcaaagtgtgcgagttccag

ttctgcaacgaccccttcctgggcgtctactatcacaagaacaacaagagctggatggaaagcgagttcc

gggtgtacagcagcgccaacaactgcaccttcgagtacgtgtcccagcctttcctgatggacctggaagg

caagcagggcaacttcaagaacctgcgcgagttcgtgttcaagaacatcgacggctacttcaagatctac

agcaagcacacccctatcaacctcgtgcgggatctgcctcagggcttctctgctctggaacccctggtgg

atctgcccatcggcatcaacatcacccggtttcagacactgctggccctgcacagaagctacctgacacc

tggcgatagcagcagcggatggacagctggtgccgccgcttactatgtgggctacctgcagcctagaacc

ttcctgctgaagtacaacgagaacggcaccatcaccgacgccgtggattgtgctctggatcctctgagcg

agacaaagtgcaccctgaagtccttcaccgtggaaaagggcatctaccagaccagcaacttccgggtgca

gcccaccgaatccatcgtgcggttccccaatatcaccaatctgtgccccttcggcgaggtgttcaatgcc

accagatcgcctctgtgtacgcctggaaccggaagcggatcagcaattgcgtggccgactactccgtgc

tgtacaactccgccagcttcagcaccttcaagtgctacggcgtgtcccctaccaagctgaacgacctgtg

cttcacaaacgtgtacgccgacagcttcgtgatccggggagatgaagtgcggcagattgcccctggacag

acaggcaacatcgccgactacaactacaagctgcccgacgacttcaccggctgtgtgattgcctggaaca

gcaacaacctggactccaaagtcggcggcaactacaattacctgtaccggctgttccggaagtccaatct

gaagcccttcgagcgggacatctccaccgagatctatcaggccggcagcaccccttgtaacggcgtgaaa

ggcttcaactgctacttcccactgcagtcctacggctttcagcccacaaatggcgtgggctatcagccct

acagagtggtggtgctgagcttcgaactgctgcatgcccctgccacagtgtgcggccctaagaaaagcac

caatctcgtgaagaacaaatgcgtgaacttcaacttcaacggcctgaccggcaccggcgtgctgacagag

agcaacaagaagttcctgccattccagcagtttggccgggatatcgccgataccacagacgccgttagag

atccccagacactggaaatcctggacatcaccccttgcagcttcggcggagtgtctgtgatcacccctgg

caccaacaccagcaatcaggtggcagtgctgtaccagggcgtgaactgtaccgaagtgcccgtggccatt

cacgccgatcagctgacacctacatggcgggtgtactccaccggcagcaatgtgtttcagaccagagccg

gctgtctgatcggagccgagcacgtgaacaatagctacgagtgcgacatccccatcggcgctggcatctg

tgccagctaccagacacagtga</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="10">

<INSDSeq>

<INSDSeq_length>1182</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..1182</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q20">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>atgttcgtgtttctggtgctgctgcctctggtgtccagccagaccctgaagt

ccttcaccgtggaaaagggcatctaccagaccagcaacttccgggtgcagcccaccgaatccatcgtgcg

gttccccaatatcaccaatctgtgccccttcggcgaggtgttcaatgccaccagattcgcctctgtgtac

gcctggaaccggaagcggatcagcaattgcgtggccgactactccgtgctgtacaactccgccagcttca

gcaccttcaagtgctacggcgtgtcccctaccaagctgaacgacctgtgcttcacaaacgtgtacgccga

cagcttcgtgatccggggagatgaagtgcggcagattgcccctggacagacaggcaacatcgccgactac

aactacaagctgcccgacgacttcaccggctgtgtgattgcctggaacagcaacaacctggactccaaag

tcggcggcaactacaattacctgtaccggctgttccggaagtccaatctgaagcccttcgagcgggacat

ctccaccgagatctatcaggccggcagcaccccttgtaacggcgtgaaaggcttcaactgctacttccca

ctgcagtcctacggctttcagcccacaaatggcgtgggctatcagccctacagagtggtggtgctgagct

tcgaactgctgcatgcccctgccacagtgtgcggccctaagaaaagcaccaatctcgtgaagaacaaatg

cgtgaacttcaacttcaacggcctgaccggcaccggcgtgctgacagagagcaacaagaagttcctgcca

ttccagcagtttggccgggatatcgccgataccacagacgccgttagagatccccagacactggaaatcc

tggacatcaccccttgcagcttcggcggagtgtctgtgatcacccctggcaccaacaccagcaatcaggt

ggcagtgctgtaccagggcgtgaactgtaccgaagtgcccgtggccattcacgccgatcagctgacacct

acatggcgggtgtactccaccggcagcaatgtgtttcagaccagagccggctgtctgatcggagccgagc

acgtgaacaatagctacgagtgcgacatccccatcggcgctggcatctgtgccagctaccagacacagac

aaacagctga</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="11">

<INSDSeq>

<INSDSeq_length>3813</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..3813</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q22">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>atgttcgtgtttctggtgctgctgcctctggtgtccagccagtgtgtgaacc

tgaccacaagaacccagctgcctccagcctacaccaacagctttaccagaggcgtgtactaccccgacaa

ggtgttcagatccagcgtgctgcactctacccaggacctgttcctgcctttcttcagcaacgtgacctgg

ttccacgccatccacgtgtccggcaccaatggcaccaagagattcgacaaccccgtgctgcccttcaacg

acggggtgtactttgccagcaccgagaagtccaacatcatcagaggctggatcttcggcaccacactgga

cagcaagacccagcctgctgatcgtgaacaacgccaccaacgtggtcatcaaagtgtgcgagttccag

ttctgcaacgaccccttcctgggcgtctactatcacaagaacaacaagagctggatggaaagcgagttcc

gggtgtacagcagcgccaacaactgcaccttcgagtacgtgtcccagcctttcctgatggacctggaagg

caagcagggcaacttcaagaacctgcgcgagttcgtgttcaagaacatcgacggctacttcaagatctac

agcaagcacacccctatcaacctcgtgcgggatctgcctcagggcttctctgctctggaacccctggtgg

atctgcccatcggcatcaacatcacccggtttcagacactgctggccctgcacagaagctacctgacacc

tggcgatagcagcagcggatggacagctggtgccgccgcttactatgtgggctacctgcagcctagaacc

ttcctgctgaagtacaacgagaacggcaccatcaccgacgccgtggattgtgctctggatcctctgagcg

agacaaagtgcaccctgaagtccttcaccgtggaaaagggcatctaccagaccagcaacttccgggtgca

gcccaccgaatccatcgtgcggttccccaatatcaccaatctgtgccccttcggcgaggtgttcaatgcc

accagatcgcctctgtgtacgcctggaaccggaagcggatcagcaattgcgtggccgactactccgtgc

tgtacaactccgccagcttcagcaccttcaagtgctacggcgtgtcccctaccaagctgaacgacctgtg

cttcacaaacgtgtacgccgacagcttcgtgatccggggagatgaagtgcggcagattgcccctggacag

acaggcaacatcgccgactacaactacaagctgcccgacgacttcaccggctgtgtgattgcctggaaca

gcaacaacctggactccaaagtcggcggcaactacaattacaggtaccggctgttccggaagtccaatct

gaagcccttcgagcgggacatctccaccgagatctatcaggccggcagcaaaccttgtaacggcgtggaa

ggcttcaactgctacttcccactgcagtcctacggctttcagcccacaaatggcgtgggctatcagccct

acagagtggtggtgctgagcttcgaactgctgcatgcccctgccacagtgtgcggccctaagaaaagcac

caatctcgtgaagaacaaatgcgtgaacttcaacttcaacggcctgaccggcaccggcgtgctgacagag

agcaacaagaagttcctgccattccagcagtttggccgggatatcgccgataccacagacgccgttagag

atccccagacactggaaatcctggacatcaccccttgcagcttcggcggagtgtctgtgatcacccctgg

caccaacaccagcaatcaggtggcagtgctgtaccagggcgtgaactgtaccgaagtgcccgtggccatt

cacgccgatcagctgacacctacatggcgggtgtactccaccggcagcaatgtgtttcagaccagagccg

gctgtctgatcggagccgagcacgtgaacaatagctacgagtgcgacatccccatcggcgctggcatctg

tgccagctaccagacacagacaaacagccctgcttctgtggccagccagagcatcattgcctacacaatg

tctctgggcgccgagaacagcgtggcctactccaacaactctatcgctatccccaccaacttcaccatca

gcgtgaccacagatcctgcctgtgtccatgaccaagaccagcgtggactgcaccatgtacatctgcgg

cgattccaccgagtgctccaacctgctgctgcagtacggcagcttctgcacccagctgaatagagccctg

acagggatcgccgtggaacaggacaagaacacccaagaggtgttcgcccaagtgaagcagatctacaaga

cccctcctatcaaggacttcggcggcttcaatttcagccagattctgcccgatcctagcaagcccagcaa

gcggagcttcatcgaggacctgctgttcaacaaagtgacactggccgacgccggcttcatcaagcagtat

ggcgattgtctgggcgacattgccgccagggatctgatttgcgcccagaagtttaacggactgacagtgc

tgccaccactgctgaccgatgagatgatcgcccagtacacatctgccctgctggccggcacaatcacaag

cggctggacatttggagctggcgccgctctgcagatcccctttgctatgcagatggcctaccggttcaac

ggcatcggagtgacccagaatgtgctgtacgagaaccagaagctgatcgccaaccagttcaacagcgcca

tcggcaagatccaggacagcctgagcagcacagcaagcgccctgggaaagctgcaggacgtggtcaacca

gaatgcccaggcactgaacaccctggtcaagcagctgtcctccaacttcggcgccatcagctctgtgctg

aacgatatcctgagcagactggaccctccggaagccgaggtgcagatcgacagactgatcaccggaaggc

tgcagtccctgcagacctacgttacccagcagctgatcagagccgccgagattagagcctctgccaatct

ggccgccaccaagatgtctgagtgtgtgctgggccagagcaagagagtggacttttgcggcaagggctac

cacctgatgagcttccctcagtctgcccctcacggcgtggtgtttctgcacgtgacatacgtgcccgctc

aagagaagaatttcaccaccgctccagccatctgccacgacggcaaagcccactttcctagagaaggcgt

gttcgtgtccaacggcacccattggttcgtgacccagcggaacttctacgagccccagatcatcaccacc

gacaacaccttcgtgtctggcaactgcgacgtcgtgatcggcattgtgaacaataccgtgtacgaccctc

tgcagcccgagctggacagcttcaaagaggaactggataagtactttaagaaccacacaagccccgacgt

ggacctgggcgatatcagcggaatcaatgccagcgtcgtgaacatccagaaagagatcgaccggctgaac

gaggtggccaagaatctgaacgagagcctgatcgacctgcaagaactggggaagtacgagcagtacatca

agtggccctggtacatctggctggggctttatcgccggactgattgccatcgtgatggtcacaatcatgct

gtgttgcatgaccagctgctgtagctgcctgaagggctgttgtagctgtggcagctgctgcaagttcgac

gaggacgattctgagcccgtgctgaagggcgtgaaactgcactacacatga</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="12">

<INSDSeq>

<INSDSeq_length>7641</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..7641</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q24">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>ggagtggccaactccatcactaggggttcctgcggccgcacgcgtttagacg

ctagactagcatgcgactcttcgcgatgtacgggccagatatacgcgttgacattgattattgactagtt

attaatagtaatcaattacggggtcattagttcatagcccatatatggagttccgcgttacataacttac

ggtaaatggcccgcctggctgaccgcccaacgacccccgcccatgacgtcaataatgacgtatgttccc

atagtaacgccaatagggactttccattgacgtcaatgggtggactatttacggtaaactgcccacttgg

cagtacatcaagtgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaatggcccgcctg

gcattatgcccagtacatgaccttatgggactttcctacttggcagtacatctacgtattagtcatcgct

attaccatggtgatgcggttttggcagtacatcaatgggcgtggatagcggtttgactcacggggatttc

caagtctccaccccattgacgtcaatgggagtttgttttggcaccaaaatcaacgggactttccaaaatg

tcgtaacaactccgccccattgacgcaaatgggcggtaggcgtgtacggtgggaggtctatataagcaga

gctctctggctaactagagaacccactgcttactggcttatcgaaattaatacgactcactataggcgag

acccaagctggctagccgccaccatgttcgtgtttctggtgctgctgcctctggtgtccagccagtgtgt

gaacctgaccacaagaacccagctgcctccagcctacaccaacagctttaccagaggcgtgtactacccc

gacaaggtgttcagatccagcgtgctgcactctacccaggacctgttcctgcctttcttcagcaacgtga

cctggttccacgccatccacgtgtccggcaccaatggcaccaagagattcgacaaccccgtgctgccctt

caacgacggggtgtactttgccagcaccgagaagtccaacatcatcagaggctggatcttcggcaccaca

ctggacagcaagacccagagcctgctgatcgtgaacaacgccaccaacgtggtcatcaaagtgtgcgagt

tccagttctgcaacgaccccttcctgggcgtctactatcacaagaacaacaagagctggatggaaagcga

gttccgggtgtacagcagcgccaacaactgcaccttcgagtacgtgtcccagcctttcctgatggacctg

gaaggcaagcagggcaacttcaagaacctgcgcgagttcgtgttcaagaacatcgacggctacttcaaga

tctacagcaagcacacccctatcaacctcgtgcgggatctgcctcagggcttctctgctctggaacccct

ggtggatctgcccatcggcatcaacatcacccggtttcagacactgctggccctgcacagaagctacctg

acacctggcgatagcagcagcggatggacagctggtgccgccgcttactatgtgggctacctgcagccta

gaaccttcctgctgaagtacaacgagaacggcaccatcaccgacgccgtggattgtgctctggatcctct

gagcgagacaaagtgcaccctgaagtccttcaccgtggaaaagggcatctaccagaccagcaacttccgg

gtgcagcccaccgaatccatcgtgcggttccccaatatcaccaatctgtgccccttcggcgaggtgttca

atgccaccagattcgcctctgtgtacgcctggaaccggaagcggatcagcaattgcgtggccgactactc

cgtgctgtacaactccgccagcttcagcaccttcaagtgctacggcgtgtcccctaccaagctgaacgac

ctgtgcttcacaaacgtgtacgccgacagcttcgtgatccggggagatgaagtgcggcagattgcccctg

gacagacaggcaagatcgccgactacaactacaagctgcccgacgacttcaccggctgtgtgattgcctg

gaacagcaacaacctggactccaaagtcggcggcaactacaattacctgtaccggctgttccggaagtcc

aatctgaagcccttcgagcgggacatctccaccgagatctatcaggccggcagcaccccttgtaacggcg

tggaaggcttcaactgctacttcccactgcagtcctacggctttcagcccacaaatggcgtgggctatca

gccctacagagtggtggtgctgagcttcgaactgctgcatgcccctgccacagtgtgcggccctaagaaa

agcaccaatctcgtgaagaacaaatgcgtgaacttcaacttcaacggcctgaccggcaccggcgtgctga

cagagagcaacaagaagttcctgccattccagcagtttggccgggatatcgccgataccacagacgccgt

tagagatccccagacactggaaatcctggacatcaccccttgcagcttcggcggagtgtctgtgatcacc

cctggcaccaacaccagcaatcaggtggcagtgctgtaccagggcgtgaactgtaccgaagtgcccgtgg

ccattcacgccgatcagctgacacctacatggcgggtgtactccaccggcagcaatgtgtttcagaccag

agccggctgtctgatcggagccgagcacgtgaacaatagctacgagtgcgacatccccatcggcgctggc

atctgtgccagctaccagacacagacaaacagccctgcttctgtggccagccagagcatcattgcctaca

caatgtctctgggcgccgagaacagcgtggcctactccaacaactctatcgctatccccaccaacttcac

catcagcgtgaccacagagatcctgcctgtgtccatgaccaagaccagcgtggactgcaccatgtacatc

tgcggcgattccaccgagtgctccaacctgctgctgcagtacggcagcttctgcacccagctgaatagag

ccctgacagggatcgccgtggaacaggacaagaacacccaagaggtgttcgcccaagtgaagcagatcta

caagacccctcctatcaaggacttcggcggcttcaatttcagccagattctgcccgatcctagcaagccc

agcaagcggagcttcatcgaggacctgctgttcaacaaagtgacactggccgacgccggcttcatcaagc

agtatggcgattgtctgggcgacattgccgccagggatctgatttgcgcccagaagtttaacggactgac

agtgctgccaccactgctgaccgatgagatgatcgcccagtacacatctgccctgctggccggcacaatc

acaagcggctggacatttggagctggcgccgctctgcagatccccttgctatgcagatggcctaccggt

tcaacggcatcggagtgacccagaatgtgctgtacgagaaccagaagctgatcgccaaccagttcaacag

cgccatcggcaagatccaggacagcctgagcagcacagcaagcgccctgggaaagctgcaggacgtggtc

aaccagaatgcccaggcactgaacaccctggtcaagcagctgtcctccaacttcggcgccatcagctctg

tgctgaacgatatcctgagcagactggaccctccggaagccgaggtgcagatcgacagactgatcaccgg

aaggctgcagtccctgcagacctacgttacccagcagctgatcagagccgccgagattagagcctctgcc

aatctggccgccaccaagatgtctgagtgtgtgctgggccagagcaagagagtggacttttgcggcaagg

gctaccacctgatgagcttccctcagtctgcccctcacggcgtggtgtttctgcacgtgacatacgtgcc

cgctcaagagaagaatttcaccaccgctccagccatctgccacgacggcaaagcccactttcctagagaa

ggcgtgttcgtgtccaacggcacccattggttcgtgacccagcggaacttctacgagccccagatcatca

ccaccgacaacaccttcgtgtctggcaactgcgacgtcgtgatcggcattgtgaacaataccgtgtacga

ccctctgcagcccgagctggacagcttcaaagaggaactggataagtactttaagaaccacacaagcccc

gacgtggacctgggcgatatcagcggaatcaatgccagcgtcgtgaacatccagaaagagatcgaccggc

tgaacgaggtggccaagaatctgaacgagagcctgatcgacctgcaagaactggggaagtacgagcagta

catcaagtggccctggtacatctggctgggctttatcgccggactgattgccatcgtgatggtcacaatc

atgctgtgttgcatgaccagctgctgtagctgcctgaagggctgttgtagctgtggcagctgctgcaagt

tcgacgaggacgattctgagcccgtgctgaagggcgtgaaactgcactacacatgagggcccgtttaaac

ccgctgatcagcctcgactgtgccttctagttgccagccatctctgttgtttgcccctcccccgtgccttcc

ttgaccctggaaggtgccactcccactgtcctttcctaataaaatgaggaaattgcatcgcattgtctga

gtaggtgtcattctattctggggggtggggtggggcaggacagcaagggggaggattgggaagacaatag

caggcatgctggggatgcggtgggctctatggcttctactgggcggttttatggacagcaaaacaacaac

aattggatctgcgctgattttgtaggtaaccacgtgcggaccgagcggccgcaggaacccctagtgatgg

agttggccactccctctctgcgcgctcgctcgctcactgaggccgggcgaccaaaggtcgcccgacgccc

gggctttgcccgggcggcctcagtgagcgagcgagcgcgcagctgcctgcaggcttggatcccaatggcg

cgccgagcttggctcgagcatggtcatagctgtttcctgtgtgaaattgttatccgctcacaattccaca

caacatacgagccggaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactcacattaatt

gcgttgcgctcactgcccgctttccagtcgggaaacctgtcgtgccagctgcattaatgaatcggccaac

gcgcggggagaggcggtttgcgtattgggcgctcttccgcttcctcgctcactgactcgctgcgctcggt

cgttcggctgcggcgagcggtatcagctcactcaaaggcggtaatacggttatccacagaatcaggggat

aacgcaggaaagaacatgtgagcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctgg

cgtttttccataggctccgcccccctgacgagcatcacaaaaatcgacgctcaagtcagaggtggcgaaa

cccgacaggactataaagataccaggcgtttccccctggaagctccctcgtgcgctctcctgttccgacc

ctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgctttctcatagctcacgct

gtaggtatctcagttcggtgtaggtcgttcgctccaagctgggctgtgtgcacgaaccccccgttcagcc

cgaccgctgcgccttatccggtaactatcgtcttgagtccaacccggtaagacacgacttatcgccactg

gcagcagccactggtaacaggattagcagagcgaggtatgtaggcggtgctacagagttcttgaagtggt

ggcctaactacggctacactagaagaacagtatttggtatctgcgctctgctgaagccagttaccttcgg

aaaaagagttggtagctcttgatccggcaaacaaaccaccgctggtagcggtggtttttttgtttgcaag

cagcagattacgcgcagaaaaaaaggatctcaagaagatcctttgatcttttctacggggtctgacgctc

agtggaacgaaaactcacgttaagggattttggtcatgagattatcaaaaaggatcttcacctagatcct

tttaaattaaaaatgaagttttaaatcaatctaaagtatatatgagtaaacttggtctgacagttagaaa

aactcatcgagcatcaaatgaaactgcaatttattcatatcaggattatcaataccatatttttgaaaaa

gccgtttctgtaatgaaggagaaaactcaccgaggcagttccataggatggcaagatcctggtatcggtc

tgcgattccgactcgtccaacatcaatacaacctattaatttcccctcgtcaaaaataaggttatcaagt

gagaaatcaccatgagtgacgactgaatccggtgagaatggcaaaagtttatgcatttctttccagactt

gttcaacaggccagccattacgctcgtcatcaaaatcactcgcatcaaccaaaccgttattcattcgtga

ttgcgcctgagcgagacgaaatacgcgatcgctgttaaaaggacaattacaaacaggaatcgaatgcaac

cggcgcaggaacactgccagcgcatcaacaatattttcacctgaatcaggatattcttctaatacctgga

atgctgttttcccagggatcgcagtggtgagtaaccatgcatcatcaggagtacggataaaatgcttgat

ggtcggaagaggcataaattccgtcagccagtttagtctgaccatctcatctgtaacatcattggcaacg

ctacctttgccatgtttcagaaacaactctggcgcatcgggcttcccatacaatcgatagattgtcgcac

ctgattgcccgacattatcgcgagcccatttatacccatataaatcagcatccatgttggaatttaatcg

cggcctagagcaagacgtttcccgttgaatatggctcatactcttcctttttcaatattattgaagcatt

tatcagggttattgtctcatgagcggatacatatttgaatgtatttagaaaaataaacaaataggggttc

cgcgcacatttccccgaaaagtgccacctgacgtctaagaaaccattattatcatgacattaacctataa

aaataggcgtatcacgaggccctttcgtctcgcgcgtttcggtgatgacggtgaaaacctctgacacatg

cagctcccggagacggtcacagcttgtctgtaagcggatgccgggagcagacaagcccgtcagggcgcgt

cagcgggtgttggcgggtgtcggggctggcttaactatgcggcatcagagcagattgtactgagagtgca

ccatatgcggtgtgaaataccgcacagatgcgtaaggagaaaataccgcatcaggcgcccctgcaggcag

ctgcgcgctcgctcgctcactgaggccgcccgggcaaagcccgggcgtcgggcgacctttggtcgcccgg

cctcagtgagcgagcgagcgcgcagagag</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="13">

<INSDSeq>

<INSDSeq_length>7640</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..7640</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q26">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>cgcccctgcaggcagctgcgcgctcgctcgctcactgaggccgcccgggcaa

agcccgggcgtcgggcgacctttggtcgcccggcctcagtgagcgagcgagcgcgcagagagggagtggc

caactccatcactaggggttcctgcggccgcacgcgtttagacgctagactagcatgcgactcttcgcga

tgtacgggccagatatacgcgttgacattgattattgactagttattaatagtaatcaattacggggtca

ttagttcatagcccatatatggagttccgcgttacataacttacggtaaatggcccgcctggctgaccgc

ccaacgacccccgcccattgacgtcaataatgacgtatgttcccatagtaacgccaatagggactttcca

ttgacgtcaatgggtggactatttacggtaaactgcccacttggcagtacatcaagtgtatcatatgcca

agtacgccccctattgacgtcaatgacggtaaatggcccgcctggcattatgcccagtacatgaccttat

gggactttcctacttggcagtacatctacgtattagtcatcgctattaccatggtgatgcggttttggca

gtacatcaatgggcgtggatagcggtttgactcacggggatttccaagtctccaccccattgacgtcaat

gggagtttgttttggcaccaaaatcaacgggactttccaaaatgtcgtaacaactccgccccattgacgc

aaatgggcggtaggcgtgtacggtgggaggtctatataagcagagctctctggctaactagagaacccac

tgcttactggcttatcgaaattaatacgactcactatagggagacccaagctggctagccgccaccatgt

tcgtgtttctggtgctgctgcctctggtgtccagccagtgtgtgaacctgaccacaagaacccagctgcc

tccagcctacaccaacagctttaccagaggcgtgtactaccccgacaaggtgttcagatccagcgtgctg

cactctacccaggacctgttcctgcctttcttcagcaacgtgacctggttccacgccatccacgtgtccg

gcaccaatggcaccaagagattcgacaaccccgtgctgcccttcaacgacggggtgtactttgccagcac

cgagaagtccaacatcatcagaggctggatcttcggcaccacactggacagcaagacccagagcctgctg

atcgtgaacaacgccaccaacgtggtcatcaaagtgtgcgagttccagttctgcaacgaccccttcctgg

gcgtctactatcacaagaacaacaagagctggatggaaagcgagttccgggtgtacagcagcgccaacaa

ctgcaccttcgagtacgtgtcccagcctttcctgatggacctggaaggcaagcagggcaacttcaagaac

ctgcgcgagttcgtgttcaagaacatcgacggctacttcaagatctacagcaagcacacccctatcaacc

tcgtgcgggatctgcctcagggcttctctgctctggaacccctggtggatctgcccatcggcatcaacat

cacccggtttcagacactgctggccctgcacagaagctacctgacacctggcgatagcagcagcggatgg

acagctggtgccgccgcttactatgtgggctacctgcagcctagaaccttcctgctgaagtacaacgaga

acggcaccatcaccgacgccgtggattgtgctctggatcctctgagcgagacaaagtgcaccctgaagtc

cttcaccgtggaaaagggcatctaccagaccagcaacttccgggtgcagcccaccgaatccatcgtgcgg

ttccccaatatcaccaatctgtgccccttcggcgaggtgttcaatgccaccagattcgcctctgtgtacg

cctggaaccggaagcggatcagcaattgcgtggccgactactccgtgctgtacaactccgccagcttcag

caccttcaagtgctacggcgtgtcccctaccaagctgaacgacctgtgcttcacaaacgtgtacgccgac

agcttcgtgatccggggagatgaagtgcggcagattgcccctggacagacaggcaacatcgccgactaca

actacaagctgcccgacgacttcaccggctgtgtgattgcctggaacagcaacaacctggactccaaagt

cggcggcaactacaattacctgtaccggctgttccggaagtccaatctgaagcccttcgagcgggacatc

tccaccgagatctatcaggccggcagcaccccttgtaacggcgtgaaaggcttcaactgctacttcccac

tgcagtcctacggctttcagcccacaaatggcgtgggctatcagccctacagagtggtggtgctgagctt

cgaactgctgcatgcccctgccacagtgtgcggccctaagaaaagcaccaatctcgtgaagaacaaatgc

gtgaacttcaacttcaacggcctgaccggcaccggcgtgctgacagagagcaacaagaagttcctgccat

tccagcagtttggccgggatatcgccgataccacagacgccgttagagatccccagacactggaaatcct

ggacatcaccccttgcagcttcggcggagtgtctgtgatcacccctggcaccaacaccagcaatcaggtg

gcagtgctgtaccagggcgtgaactgtaccgaagtgcccgtggccattcacgccgatcagctgacaccta

catggcgggtgtactccaccggcagcaatgtgtttcagaccagagccggctgtctgatcggagccgagca

cgtgaacaatagctacgagtgcgacatccccatcggcgctggcatctgtgccagctaccagacacagaca

aacagccctgcttctgtggccagccagagcatcattgcctacacaatgtctctgggcgccgagaacagcg

tggcctactccaacaactctatcgctatccccaccaacttcaccatcagcgtgaccacagagatcctgcc

tgtgtccatgaccaagaccagcgtggactgcaccatgtacatctgcggcgattccaccgagtgctccaac

ctgctgctgcagtacggcagcttctgcacccagctgaatagagccctgacagggatcgccgtggaacagg

acaagaacacccaagaggtgttcgcccaagtgaagcagatctacaagacccctcctatcaaggacttcgg

cggcttcaatttcagccagattctgcccgatcctagcaagcccagcaagcggagcttcatcgaggacctg

ctgttcaacaaagtgacactggccgacgccggcttcatcaagcagtatggcgattgtctgggcgacattg

ccgccagggatctgatttgcgcccagaagtttaacggactgacagtgctgccaccactgctgaccgatga

gatgatcgcccagtacacatctgccctgctggccggcacaatcacaagcggctggacatttggagctggc

gccgctctgcagatcccctttgctatgcagatggcctaccggttcaacggcatcggagtgacccagaatg

tgctgtacgagaaccagaagctgatcgccaaccagttcaacagcgccatcggcaagatccaggacagcct

gagcagcacagcaagcgccctgggaaagctgcaggacgtggtcaaccagaatgcccaggcactgaacacc

ctggtcaagcagctgtcctccaacttcggcgccatcagctctgtgctgaacgatatcctgagcagactgg

accctccggaagccgaggtgcagatcgacagactgatcaccggaaggctgcagtccctgcagacctacgt

tacccagcagctgatcagagccgccgagattagagcctctgccaatctggccgccaccaagatgtctgag

tgtgtgctgggccagagcaagagagtggacttttgcggcaagggctaccacctgatgagcttccctcagt

ctgcccctcacggcgtggtgtttctgcacgtgacatacgtgcccgctcaagagaagaatttcaccaccgc

tccagccatctgccacgacggcaaagcccactttcctagagaaggcgtgttcgtgtccaacggcacccat

tggttcgtgacccagcggaacttctacgagccccagatcatcaccaccgacaacaccttcgtgtctggca

actgcgacgtcgtgatcggcattgtgaacaataccgtgtacgaccctctgcagcccgagctggacagctt

caaagaggaactggataagtactttaagaaccacacaagccccgacgtggacctgggcgatatcagcgga

atcaatgccagcgtcgtgaacatccagaaagagatcgaccggctgaacgaggtggccaagaatctgaacg

agagcctgatcgacctgcaagaactggggaagtacgagcagtacatcaagtggccctggtacatctggct

gggctttatcgccggactgattgccatcgtgatggtcacaatcatgctgtgttgcatgaccagctgctgt

agctgcctgaagggctgttgtagctgtggcagctgctgcaagttcgacgaggacgattctgagcccgtgc

tgaagggcgtgaaactgcactacacatgagggcccgtttaaacccgctgatcagcctcgactgtgccttc

tagttgccagccatctgttgtttgcccctcccccgtgccttccttgaccctggaaggtgccactcccact

gtcctttcctaataaaatgaggaaattgcatcgcattgtctgagtaggtgtcattctattctggggggtg

gggtggggcaggacagcaagggggaggattgggaagacaatagcaggcatgctggggatgcggtgggctc

tatggcttctactgggcggttttatggacagcaaaacaacaacaattggatctgcgctgattttgtaggt

aaccacgtgcggaccgagcggccgcaggaacccctagtgatggagttggccactccctctctgcgcgctc

gctcgctcactgaggccgggcgaccaaaggtcgcccgacgcccgggctttgcccgggcggcctcagtgag

cgagcgagcgcgcagctgcctgcaggcttggatcccaatggcgcgccgagcttggctcgagcatggtcat

agctgtttcctgtgtgaaattgttatccgctcacaattccacacaacatacgagccggaagcataaagtg

taaagcctggggtgcctaatgagtgagctaactcacattaattgcgttgcgctcactgcccgctttccag

tcgggaaacctgtcgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggtttgcgtattg

ggcgctcttccgcttcctcgctcactgactcgctgcgctcggtcgttcggctgcggcgagcggtatcagc

tcactcaaaggcggtaatacggttatccacagaatcaggggataacgcaggaaagaacatgtgagcaaaa

ggccagcaaaaggccaggaaccgtaaaaaggccgcgttgctggcgtttttccataggctccgcccccctg

acgagcatcacaaaaatcgacgctcaagtcagaggtggcgaaacccgacaggactataaagataccaggc

gtttccccctggaagctccctcgtgcgctctcctgttccgaccctgccgcttaccggatacctgtccgcc

tttctcccttcgggaagcgtggcgctttctcatagctcacgctgtaggtatctcagttcggtgtaggtcg

ttcgctccaagctgggctgtgtgcacgaaccccccgttcagcccgaccgctgcgccttatccggtaacta

tcgtcttgagtccaacccggtaagacacgacttatcgccactggcagcagccactggtaacaggattagc

agagcgaggtatgtaggcggtgctacagagttcttgaagtggtggcctaactacggctacactagaagaa

cagtatttggtatctgcgctctgctgaagccagttaccttcggaaaaagagttggtagctcttgatccgg

caaacaaaccaccgctggtagcggtggtttttttgtttgcaagcagcagattacgcgcagaaaaaaagga

tctcaagaagatcctttgatcttttctacggggtctgacgctcagtggaacgaaaactcacgttaaggga

ttttggtcatgagattatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaatc

aatctaaagtatatgagtaaacttggtctgacagttagaaaaactcatcgagcatcaaatgaaactgc

aatttattcatatcaggattatcaataccatatttttgaaaaagccgtttctgtaatgaaggagaaaact

caccgaggcagttccataggatggcaagatcctggtatcggtctgcgattccgactcgtccaacatcaat

acaacctattaatttcccctcgtcaaaaataaggttatcaagtgagaaatcaccatgagtgacgactgaa

tccggtgagaatggcaaaagtttatgcatttctttccagacttgttcaacaggccagccattacgctcgt

catcaaaatcactcgcatcaaccaaaccgttattcattcgtgattgcgcctgagcgagacgaaatacgcg

atcgctgttaaaaggacaattacaaacaggaatcgaatgcaaccggcgcaggaacactgccagcgcatca

acaatattttcacctgaatcaggatattcttctaatacctggaatgctgttttcccagggatcgcagtgg

tgagtaaccatgcatcatcaggagtacggataaaatgcttgatggtcggaagaggcataaattccgtcag

ccagtttagtctgaccatctcatctgtaacatcattggcaacgctacctttgccatgtttcagaaacaac

tctggcgcatcgggcttcccatacaatcgatagattgtcgcacctgattgcccgacattatcgcgagccc

atttatacccatataaatcagcatccatgttggaatttaatcgcggcctagagcaagacgtttcccgttg

aatatggctcatactcttcctttttcaatattattgaagcatttatcagggttattgtctcatgagcgga

tacatatttgaatgtatttagaaaaataaacaaataggggttccgcgcacatttccccgaaaagtgccac

ctgacgtctaagaaaccattattatcatgacattaacctataaaaataggcgtatcacgaggccctttcg

tctcgcgcgtttcggtgatgacggtgaaaacctctgacacatgcagctcccggagacggtcacagcttgt

ctgtaagcggatgccgggagcagacaagcccgtcagggcgcgtcagcgggtgttggcgggtgtcggggct

ggcttaactatgcggcatcagagcagattgtactgagagtgcaccatatgcggtgtgaaataccgcacag

atgcgtaaggagaaaataccgcatcagg</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="14">

<INSDSeq>

<INSDSeq_length>5861</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..5861</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q28">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>cgcccctgcaggcagctgcgcgctcgctcgctcactgaggccgcccgggcaa

agcccgggcgtcgggcgacctttggtcgcccggcctcagtgagcgagcgagcgcgcagagagggagtggc

caactccatcactaggggttcctgcggccgcacgcgtttagacgctagactagcatgcgactcttcgcga

tgtacgggccagatatacgcgttgacattgattattgactagttattaatagtaatcaattacggggtca

ttagttcatagcccatatatggagttccgcgttacataacttacggtaaatggcccgcctggctgaccgc

ccaacgacccccgcccattgacgtcaataatgacgtatgttcccatagtaacgccaatagggactttcca

ttgacgtcaatgggtggactatttacggtaaactgcccacttggcagtacatcaagtgtatcatatgcca

agtacgccccctattgacgtcaatgacggtaaatggcccgcctggcattatgcccagtacatgaccttat

gggactttcctacttggcagtacatctacgtattagtcatcgctattaccatggtgatgcggttttggca

gtacatcaatgggcgtggatagcggtttgactcacggggatttccaagtctccaccccattgacgtcaat

gggagtttgttttggcaccaaaatcaacgggactttccaaaatgtcgtaacaactccgccccattgacgc

aaatgggcggtaggcgtgtacggtgggaggtctatataagcagagctctctggctaactagagaacccac

tgcttactggcttatcgaaattaatacgactcactatagggagacccaagctggctagccgccaccatgt

tcgtgtttctggtgctgctgcctctggtgtccagccagtgtgtgaacctgaccacaagaacccagctgcc

tccagcctacaccaacagctttaccagaggcgtgtactaccccgacaaggtgttcagatccagcgtgctg

cactctacccaggacctgttcctgcctttcttcagcaacgtgacctggttccacgccatccacgtgtccg

gcaccaatggcaccaagagattcgacaaccccgtgctgcccttcaacgacggggtgtactttgccagcac

cgagaagtccaacatcatcagaggctggatcttcggcaccacactggacagcaagacccagagcctgctg

atcgtgaacaacgccaccaacgtggtcatcaaagtgtgcgagttccagttctgcaacgaccccttcctgg

gcgtctactatcacaagaacaacaagagctggatggaaagcgagttccgggtgtacagcagcgccaacaa

ctgcaccttcgagtacgtgtcccagcctttcctgatggacctggaaggcaagcagggcaacttcaagaac

ctgcgcgagttcgtgttcaagaacatcgacggctacttcaagatctacagcaagcacacccctatcaacc

tcgtgcgggatctgcctcagggcttctctgctctggaacccctggtggatctgcccatcggcatcaacat

cacccggtttcagacactgctggccctgcacagaagctacctgacacctggcgatagcagcagcggatgg

acagctggtgccgccgcttactatgtgggctacctgcagcctagaaccttcctgctgaagtacaacgaga

acggcaccatcaccgacgccgtggattgtgctctggatcctctgagcgagacaaagtgcaccctgaagtc

cttcaccgtggaaaagggcatctaccagaccagcaacttccgggtgcagcccaccgaatccatcgtgcgg

ttccccaatatcaccaatctgtgccccttcggcgaggtgttcaatgccaccagattcgcctctgtgtacg

cctggaaccggaagcggatcagcaattgcgtggccgactactccgtgctgtacaactccgccagcttcag

caccttcaagtgctacggcgtgtcccctaccaagctgaacgacctgtgcttcacaaacgtgtacgccgac

agcttcgtgatccggggagatgaagtgcggcagattgcccctggacagacaggcaacatcgccgactaca

actacaagctgcccgacgacttcaccggctgtgtgattgcctggaacagcaacaacctggactccaaagt

cggcggcaactacaattacctgtaccggctgttccggaagtccaatctgaagcccttcgagcgggacatc

tccaccgagatctatcaggccggcagcaccccttgtaacggcgtgaaaggcttcaactgctacttcccac

tgcagtcctacggctttcagcccacaaatggcgtgggctatcagccctacagagtggtggtgctgagctt

cgaactgctgcatgcccctgccacagtgtgcggccctaagaaaagcaccaatctcgtgaagaacaaatgc

gtgaacttcaacttcaacggcctgaccggcaccggcgtgctgacagagagcaacaagaagttcctgccat

tccagcagtttggccgggatatcgccgataccacagacgccgttagagatccccagacactggaaatcct

ggacatcaccccttgcagcttcggcggagtgtctgtgatcacccctggcaccaacaccagcaatcaggtg

gcagtgctgtaccagggcgtgaactgtaccgaagtgcccgtggccattcacgccgatcagctgacaccta

catggcgggtgtactccaccggcagcaatgtgtttcagaccagagccggctgtctgatcggagccgagca

cgtgaacaatagctacgagtgcgacatccccatcggcgctggcatctgtgccagctaccagacacagtga

gggcccgtttaaacccgctgatcagcctcgactgtgccttctagttgccagccatctgttgtttgcccct

cccccgtgccttccttgaccctggaaggtgccactcccactgtcctttcctaataaaatgaggaaattgc

atcgcattgtctgagtaggtgtcattctattctggggggtggggtggggcaggacagcaagggggaggat

tgggaagacaatagcaggcatgctggggatgcggtgggctctatggcttctactgggcggttttatggac

agcaaaacaacaacaattggatctgcgctgattttgtaggtaaccacgtgcggaccgagcggccgcagga

acccctagtgatggagttggccactccctctctgcgcgctcgctcgctcactgaggccgggcgaccaaag

gtcgcccgacgcccgggctttgcccgggcggcctcagtgagcgagcgagcgcgcagctgcctgcaggctt

ggatcccaatggcgcgccgagcttggctcgagcatggtcatagctgtttcctgtgtgaaattgttatccg

ctcacaattccacacaacatacgagccggaagcataaagtgtaaagcctggggtgcctaatgagtgagct

aactcacattaattgcgttgcgctcactgcccgctttccagtcgggaaacctgtcgtgccagctgcatta

atgaatcggccaacgcgcggggagaggcggtttgcgtattgggcgctcttccgcttcctcgctcactgac

tcgctgcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaatacggttatcca

cagaatcaggggataacgcaggaaagaacatgtgagcaaaaggccagcaaaaggccaggaaccgtaaaaa

ggccgcgttgctggcgtttttccataggctccgcccccctgacgagcatcacaaaaatcgacgctcaagt

cagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctggaagctccctcgtgcgct

ctcctgttccgaccctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgctttc

tcatagctcacgctgtaggtatctcagttcggtgtaggtcgttcgctccaagctgggctgtgtgcacgaa

ccccccgttcagcccgaccgctgcgccttatccggtaactatcgtcttgagtccaacccggtaagacacg

acttatcgccactggcagcagccactggtaacaggattagcagagcgaggtatgtaggcggtgctacaga

gttcttgaagtggtggcctaactacggctacactagaagaacagtatttggtatctgcgctctgctgaag

ccagttaccttcggaaaaagagttggtagctctcttgatccggcaaacaaaccaccgctggtagcggtggtt

tttttgtttgcaagcagcagattacgcgcagaaaaaaaggatctcaagaagatcctttgatcttttctac

ggggtctgacgctcagtggaacgaaaactcacgttaagggattttggtcatgagattatcaaaaaggatc

ttcacctagatccttttaaattaaaaatgaagttttaaatcaatctaaagtatatatgagtaaacttggt

ctgacagttagaaaaactcatcgagcatcaaatgaaactgcaatttattcatatcaggattatcaatacc

atatttttgaaaaagccgtttctgtaatgaaggagaaaactcaccgaggcagttccataggatggcaaga

tcctggtatcggtctgcgattccgactcgtccaacatcaatacaacctattaatttcccctcgtcaaaaa

taaggttatcaagtgagaaatcaccatgagtgacgactgaatccggtgagaatggcaaaagtttatgcat

ttctttccagacttgttcaacaggccagccattacgctcgtcatcaaaatcactcgcatcaaccaaaccg

ttattcattcgtgattgcgcctgagcgagacgaaatacgcgatcgctgttaaaaggacaattacaaacag

gaatcgaatgcaaccggcgcaggaacactgccagcgcatcaacaatattttcacctgaatcaggatattc

ttctaatacctggaatgctgttttcccagggatcgcagtggtgagtaaccatgcatcatcaggagtacgg

ataaaatgcttgatggtcggaagaggcataaattccgtcagccagtttagtctgaccatctcatctgtaa

catcattggcaacgctacctttgccatgtttcagaaacaactctggcgcatcgggcttcccatacaatcg

atagattgtcgcacctgattgcccgacattatcgcgagcccatttatacccatataaatcagcatccatg

ttggaatttaatcgcggcctagagcaagacgtttcccgttgaatatggctcatactcttccttttttcaat

attattgaagcatttatcagggttattgtctcatgagcggatacatatttgaatgtatttagaaaaataa

acaaataggggttccgcgcacatttccccgaaaagtgccacctgacgtctaagaaaccattattatcatg

acattaacctataaaaataggcgtatcacgaggccctttcgtctcgcgcgtttcggtgatgacggtgaaa

acctctgacacatgcagctcccggagacggtcacagcttgtctgtaagcggatgccgggagcagacaagc

ccgtcagggcgcgtcagcgggtgttggcgggtgtcggggctggcttaactatgcggcatcagagcagatt

gtactgagagtgcaccatatgcggtgtgaaataccgcacagatgcgtaaggagaaaataccgcatcagg<

/INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="15">

<INSDSeq>

<INSDSeq_length>5009</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..5009</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q30">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>cgcccctgcaggcagctgcgcgctcgctcgctcactgaggccgcccgggcaa

agcccgggcgtcgggcgacctttggtcgcccggcctcagtgagcgagcgagcgcgcagagagggagtggc

caactccatcactaggggttcctgcggccgcacgcgtttagacgctagactagcatgcgactcttcgcga

tgtacgggccagatatacgcgttgacattgattattgactagttattaatagtaatcaattacggggtca

ttagttcatagcccatatatggagttccgcgttacataacttacggtaaatggcccgcctggctgaccgc

ccaacgacccccgcccattgacgtcaataatgacgtatgttcccatagtaacgccaatagggactttcca

ttgacgtcaatgggtggactatttacggtaaactgcccacttggcagtacatcaagtgtatcatatgcca

agtacgccccctattgacgtcaatgacggtaaatggcccgcctggcattatgcccagtacatgaccttat

gggactttcctacttggcagtacatctacgtattagtcatcgctattaccatggtgatgcggttttggca

gtacatcaatgggcgtggatagcggtttgactcacggggatttccaagtctccaccccattgacgtcaat

gggagtttgttttggcaccaaaatcaacgggactttccaaaatgtcgtaacaactccgccccattgacgc

aaatgggcggtaggcgtgtacggtgggaggtctatataagcagagctctctggctaactagagaacccac

tgcttactggcttatcgaaattaatacgactcactatagggagacccaagctggctagccgccaccatgt

tcgtgtttctggtgctgctgcctctggtgtccagccagaccctgaagtccttcaccgtggaaaagggcat

ctaccagaccagcaacttccggggtgcagcccaccgaatccatcgtgcggttccccaatatcaccaatctg

tgccccttcggcgaggtgttcaatgccaccagattcgcctctgtgtacgcctggaaccggaagcggatca

gcaattgcgtggccgactactccgtgctgtacaactccgccagcttcagcaccttcaagtgctacggcgt

gtcccctaccaagctgaacgacctgtgcttcacaaacgtgtacgccgacagcttcgtgatccggggagat

gaagtgcggcagattgcccctggacagacaggcaacatcgccgactacaactacaagctgcccgacgact

tcaccggctgtgtgattgcctggaacagcaacaacctggactccaaagtcggcggcaactacaattacct

gtaccggctgttccggaagtccaatctgaagcccttcgagcgggacatctccaccgagatctatcaggcc

ggcagcaccccttgtaacggcgtgaaaggcttcaactgctacttcccactgcagtcctacggctttcagc

ccacaaatggcgtgggctatcagccctacagagtggtggtgctgagcttcgaactgctgcatgcccctgc

cacagtgtgcggccctaagaaaagcaccaatctcgtgaagaacaaatgcgtgaacttcaacttcaacggc

ctgaccggcaccggcgtgctgacagagagcaacaagaagttcctgccattccagcagtttggccgggata

tcgccgataccacagacgccgttagagatccccagacactggaaatcctggacatcaccccttgcagctt

cggcggagtgtctgtgatcacccctggcaccaacaccagcaatcaggtggcagtgctgtaccagggcgtg

aactgtaccgaagtgcccgtggccattcacgccgatcagctgacacctacatggcgggtgtactccaccg

gcagcaatgtgtttcagaccagagccggctgtctgatcggagccgagcacgtgaacaatagctacgagtg

cgacatccccatcggcgctggcatctgtgccagctaccagacacagacaaacagctgagggcccgtttaa

acccgctgatcagcctcgactgtgccttctagttgccagccatctgttgtttgcccctcccccgtgcctt

ccttgaccctggaaggtgccactcccactgtcctttcctaataaaatgaggaaattgcatcgcattgtct

gagtaggtgtcattctattctggggggtggggtggggcaggacagcaagggggaggattgggaagacaat

agcaggcatgctggggatgcggtgggctctatggcttctactgggcggttttatggacagcaaaacaaca

acaattggatctgcgctgattttgtaggtaaccacgtgcggaccgagcggccgcaggaacccctagtgat

ggagttggccactccctctctgcgcgctcgctcgctcactgaggccgggcgaccaaaggtcgcccgacgc

ccgggctttgcccgggcggcctcagtgagcgagcgagcgcgcagctgcctgcaggcttggatcccaatgg

cgcgccgagcttggctcgagcatggtcatagctgtttcctgtgtgaaattgttatccgctcacaattcca

cacaacatacgagccggaagcataaagtgtaaagcctggggtgcctaatgagtgagctaactcacattaa

ttgcgttgcgctcactgcccgctttccagtcgggaaacctgtcgtgccagctgcattaatgaatcggcca

acgcgcggggagaggcggtttgcgtattgggcgctcttccgcttcctcgctcactgactcgctgcgctcg

gtcgttcggctgcggcgagcggtatcagctcactcaaaggcggtaatacggttatccacagaatcagggg

ataacgcaggaaagaacatgtgagcaaaaggccagcaaaaggccaggaaccgtaaaaaggccgcgttgct

ggcgtttttccataggctccgcccccctgacgagcatcacaaaaatcgacgctcaagtcagaggtggcga

aacccgacaggactataaagataccaggcgtttccccctggaagctccctcgtgcgctctcctgttccga

ccctgccgcttaccggatacctgtccgcctttctcccttcgggaagcgtggcgctttctcatagctcacg

ctgtaggtatctcagttcggtgtaggtcgttcgctccaagctgggctgtgtgcacgaaccccccgttcag

cccgaccgctgcgccttatccggtaactatcgtcttgagtccaacccggtaagacacgacttatcgccac

tggcagcagccactggtaacaggattagcagagcgaggtatgtaggcggtgctacagagttcttgaagtg

gtggcctaactacggctacactagaagaacagtatttggtatctgcgctctgctgaagccagttaccttc

ggaaaaagagttggtagctcttgatccggcaaacaaaccaccgctggtagcggtggtttttttgtttgca

agcagcagattacgcgcagaaaaaaaggatctcaagaagatcctttgatcttttctacggggtctgacgc

tcagtggaacgaaaactcacgttaagggattttggtcatgagattatcaaaaaggatcttcacctagatc

cttttaaattaaaaatgaagttttaaatcaatctaaagtatatgagtaaacttggtctgacagttaga

aaaactcatcgagcatcaaatgaaactgcaatttattcatatcaggattatcaataccatatttttgaaa

aagccgtttctgtaatgaaggagaaaactcaccgaggcagttccataggatggcaagatcctggtatcgg

tctgcgattccgactcgtccaacatcaatacaacctattaatttcccctcgtcaaaaataaggttatcaa

gtgagaaatcaccatgagtgacgactgaatccggtgagaatggcaaaagtttatgcatttctttccagac

ttgttcaacaggccagccattacgctcgtcatcaaaatcactcgcatcaaccaaaccgttattcattcgt

gattgcgcctgagcgagacgaaatacgcgatcgctgttaaaaggacaattacaaacaggaatcgaatgca

accggcgcaggaacactgccagcgcatcaacaatattttcacctgaatcaggatattcttctaatacctg

gaatgctgttttcccagggatcgcagtggtgagtaaccatgcatcatcaggagtacggataaaatgcttg

atggtcggaagaggcataaattccgtcagccagtttagtctgaccatctcatctgtaacatcattggcaa

cgctacctttgccatgtttcagaaacaactctggcgcatcgggcttcccatacaatcgatagattgtcgc

acctgattgcccgacattatcgcgagcccatttatacccatataaatcagcatccatgttggaatttaat

cgcggcctagagcaagacgtttcccgttgaatatggctcatactcttcctttttcaatattattgaagca

tttatcagggttattgtctcatgagcggatacatatttgaatgtatttagaaaaataaacaaataggggt

tccgcgcacatttccccgaaaagtgccacctgacgtctaagaaaccattattatcatgacattaacctat

aaaaataggcgtatcacgaggccctttcgtctcgcgcgtttcggtgatgacggtgaaaacctctgacaca

tgcagctcccggagacggtcacagcttgtctgtaagcggatgccgggagcagacaagcccgtcagggcgc

gtcagcgggtgttggcgggtgtcggggctggcttaactatgcggcatcagagcagattgtactgagagtg

caccatatgcggtgtgaaataccgcacagatgcgtaaggagaaaataccgcatcagg</INSDSeq_seq

uence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="16">

<INSDSeq>

<INSDSeq_length>7626</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..7626</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q32">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>cgcccctgcaggcagctgcgcgctcgctcgctcactgaggccgcccgggcaa

agcccgggcgtcgggcgacctttggtcgcccggcctcagtgagcgagcgagcgcgcagagagggagtggc

caactccatcactaggggttcctgcggccgcacgcgtttagacggactctcttcgcgatgtacgggccagat

atacgcgttgacattgattattgactagttattaatagtaatcaattacggggtcattagttcatagccc

atatatggagttccgcgttacataacttacggtaaatggcccgcctggctgaccgcccaacgacccccgc

ccattgacgtcaataatgacgtatgttcccatagtaacgccaatagggactttccattgacgtcaatggg

tggactatttacggtaaactgcccacttggcagtacatcaagtgtatcatatgccaagtacgccccctat

tgacgtcaatgacggtaaatggcccgcctggcattatgcccagtacatgaccttatgggactttcctact

tggcagtacatctacgtattagtcatcgctattaccatggtgatgcggttttggcagtacatcaatgggc

gtggatagcggtttgactcacggggatttccaagtctccaccccattgacgtcaatgggagtttgttttg

gcaccaaaatcaacgggactttccaaaatgtcgtaacaactccgccccattgacgcaaatgggcggtagg

cgtgtacggtgggaggtctatataagcagagctctctggctaactagagaacccactgcttactggctta

tcgaaattaatacgactcactatagggagacccaagctggctagccgccaccatgttcgtgtttctggtg

ctgctgcctctggtgtccagccagtgtgtgaacctgaccacaagaacccagctgcctccagcctacacca

acagctttaccagaggcgtgtactaccccgacaaggtgttcagatccagcgtgctgcactctacccagga

cctgttcctgcctttcttcagcaacgtgacctggttccacgccatccacgtgtccggcaccaatggcacc

aagagattcgacaaccccgtgctgcccttcaacgacggggtgtactttgccagcaccgagaagtccaaca

tcatcagaggctggatcttcggcaccacactggacagcaagacccagagcctgctgatcgtgaacaacgc

caccaacgtggtcatcaaagtgtgcgagttccagttctgcaacgaccccttcctgggcgtctactatcac

aagaacaacaagagctggatggaaagcgagttccgggtgtacagcagcgccaacaactgcaccttcgagt

acgtgtcccagcctttcctgatggacctggaaggcaagcagggcaacttcaagaacctgcgcgagttcgt

gttcaagaacatcgacggctacttcaagatctacagcaagcacacccctatcaacctcgtgcgggatctg

cctcagggcttctctgctctggaacccctggtggatctgcccatcggcatcaacatcacccggtttcaga

cactgctggccctgcacagaagctacctgacacctggcgatagcagcagcggatggacagctggtgccgc

cgcttactatgtgggctacctgcagcctagaaccttcctgctgaagtacaacgagaacggcaccatcacc

gacgccgtggattgtgctctggatcctctgagcgagacaaagtgcaccctgaagtccttcaccgtggaaa

agggcatctaccagaccagcaacttccgggtgcagcccaccgaatccatcgtgcggttccccaatatcac

caatctgtgccccttcggcgaggtgttcaatgccaccagattcgcctctgtgtacgcctggaaccggaag

cggatcagcaattgcgtggccgactactccgtgctgtacaactccgccagcttcagcaccttcaagtgct

acggcgtgtcccctaccaagctgaacgacctgtgcttcacaaacgtgtacgccgacagcttcgtgatccg

ggggagatgaagtgcggcagattgcccctggacagacaggcaacatcgccgactacaactacaagctgccc

gacgacttcaccggctgtgtgattgcctggaacagcaacaacctggactccaaagtcggcggcaactaca

attacaggtaccggctgttccggaagtccaatctgaagcccttcgagcgggacatctccaccgagatcta

tcaggccggcagcaaaccttgtaacggcgtggaaggcttcaactgctacttcccactgcagtcctacggc

tttcagcccacaaatggcgtgggctatcagccctacagagtggtggtgctgagcttcgaactgctgcatg

cccctgccacagtgtgcggccctaagaaaagcaccaatctcgtgaagaacaaatgcgtgaacttcaactt

caacggcctgaccggcaccggcgtgctgacagagagcaacaagaagttcctgccattccagcagtttggc

cgggatatcgccgataccacagacgccgttagagatccccagacactggaaatcctggacatcacccctt

gcagcttcggcggagtgtctgtgatcacccctggcaccaacaccagcaatcaggtggcagtgctgtacca

gggcgtgaactgtaccgaagtgcccgtggccattcacgccgatcagctgacacctacatggcgggtgtac

tccaccggcagcaatgtgtttcagaccagagccggctgtctgatcggagccgagcacgtgaacaatagct

acgagtgcgacatccccatcggcgctggcatctgtgccagctaccagacacagacaaacagccctgcttc

tgtggccagccagagcatcattgcctacacaatgtctctgggcgccgagaacagcgtggcctactccaac

aactctatcgctatccccaccaacttcaccatcagcgtgaccacagatcctgcctgtgtccatgacca

agaccagcgtggactgcaccatgtacatctgcggcgattccaccgagtgctccaacctgctgctgcagta

cggcagcttctgcacccagctgaatagagccctgacagggatcgccgtggaacaggacaagaacacccaa

gaggtgttcgcccaagtgaagcagatctacaagacccctcctatcaaggacttcggcggcttcaatttca

gccagattctgcccgatcctagcaagcccagcaagcggagcttcatcgaggacctgctgttcaacaaagt

gacactggccgacgccggcttcatcaagcagtatggcgattgtctgggcgacattgccgccagggatctg

atttgcgcccagaagtttaacggactgacagtgctgccaccactgctgaccgatgagatgatcgcccagt

acacatctgccctgctggccggcacaatcacaagcggctggacatttggagctggcgccgctctgcagat

cccctttgctatgcagatggcctaccggttcaacggcatcggagtgacccagaatgtgctgtacgagaac

cagaagctgatcgccaaccagttcaacagcgccatcggcaagatccaggacagcctgagcagcacagcaa

gcgccctgggaaagctgcaggacgtggtcaaccagaatgcccaggcactgaacaccctggtcaagcagct

gtcctccaacttcggcgccatcagctctgtgctgaacgatatcctgagcagactggaccctccggaagcc

gaggtgcagatcgacagactgatcaccggaaggctgcagtccctgcagacctacgttacccagcagctga

tcagagccgccgagattagagcctctgccaatctggccgccaccaagatgtctgagtgtgtgctgggcca

gagcaagagagtggacttttgcggcaagggctaccacctgatgagcttccctcagtctgcccctcacggc

gtggtgtttctgcacgtgacatacgtgcccgctcaagagaagaatttcaccaccgctccagccatctgcc

acgacggcaaagcccactttcctagagaaggcgtgttcgtgtccaacggcacccattggttcgtgaccca

gcggaacttctacgagccccagatcatcaccaccgacaacaccttcgtgtctggcaactgcgacgtcgtg

atcggcattgtgaacaataccgtgtacgaccctctgcagcccgagctggacagcttcaaagaggaactgg

ataagtactttaagaaccacacaagccccgacgtggacctgggcgatatcagcggaatcaatgccagcgt

cgtgaacatccagaaagagatcgaccggctgaacgaggtggccaagaatctgaacgagagcctgatcgac

ctgcaagaactggggaagtacgagcagtacatcaagtggccctggtacatctggctgggctttatcgccg

gactgattgccatcgtgatggtcacaatcatgctgtgttgcatgaccagctgctgtagctgcctgaaggg

ctgttgtagctgtggcagctgctgcaagttcgacgaggacgattctgagcccgtgctgaagggcgtgaaa

ctgcactacacatgagggcccgtttaaacccgctgatcagcctcgactgtgccttctagttgccagccat

ctgttgtttgcccctcccccgtgccttccttgaccctggaaggtgccactcccactgtcctttcctaata

aaatgaggaaattgcatcgcattgtctgagtaggtgtcattctattctggggggtggggtggggcaggac

agcaagggggaggattgggaagacaatagcaggcatgctggggatgcggtgggctctatggcttctactg

ggcggttttatggacagcaaaacaacaacaattggatctgcgctgattttgtaggtaaccacgtgcggac

cgagcggccgcaggaacccctagtgatggagttggccactccctctctgcgcgctcgctcgctcactgag

gccgggcgaccaaaggtcgcccgacgcccgggctttgcccgggcggcctcagtgagcgagcgagcgcgca

gctgcctgcaggcttggatcccaatggcgcgccgagcttggctcgagcatggtcatagctgtttcctgtg

tgaaattgttatccgctcacaattccacacaacatacgagccggaagcataaagtgtaaagcctggggtg

cctaatgagtgagctaactcacattaattgcgttgcgctcactgcccgctttccagtcgggaaacctgtc

gtgccagctgcattaatgaatcggccaacgcgcggggagaggcggtttgcgtattgggcgctcttccgct

tcctcgctcactgactcgctgcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaaggcgg

taatacggttatccacagaatcaggggataacgcaggaaagaacatgtgagcaaaaggccagcaaaaggc

caggaaccgtaaaaaggccgcgttgctggcgtttttccataggctccgcccccctgacgagcatcacaaa

aatcgacgctcaagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctggaa

gctccctcgtgcgctctcctgttccgaccctgccgcttaccggatacctgtccgcctttctcccttcggg

aagcgtggcgctttctcatagctcacgctgtaggtatctcagttcggtgtaggtcgttcgctccaagctg

ggctgtgtgcacgaaccccccgttcagcccgaccgctgcgccttatccggtaactatcgtcttgagtcca

acccggtaagacacgacttatcgccactggcagcagccactggtaacaggattagcagagcgaggtatgt

aggcggtgctacagagttcttgaagtggtggcctaactacggctacactagaagaacagtatttggtatc

tgcgctctgctgaagccagttaccttcggaaaaagagttggtagctcttgatccggcaaacaaaccaccg

ctggtagcggtggtttttttgtttgcaagcagcagattacgcgcagaaaaaaaaggatctcaagaagatcc

tttgatcttttctacggggtctgacgctcagtggaacgaaaactcacgttaagggattttggtcatgaga

ttatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaatcaatctaaagtatat

atgagtaaacttggtctgacagttagaaaaactcatcgagcatcaaatgaaactgcaatttattcatatc

aggattatcaataccatatttttgaaaaagccgtttctgtaatgaaggagaaaactcaccgaggcagttc

cataggatggcaagatcctggtatcggtctgcgattccgactcgtccaacatcaatacaacctattaatt

tcccctcgtcaaaaataaggttatcaagtgagaaatcaccatgagtgacgactgaatccggtgagaatgg

caaaagtttatgcatttctttccagacttgttcaacaggccagccattacgctcgtcatcaaaatcactc

gcatcaaccaaaccgttattcattcgtgattgcgcctgagcgagacgaaatacgcgatcgctgttaaaag

gacaattacaaacaggaatcgaatgcaaccggcgcaggaacactgccagcgcatcaacaatattttcacc

tgaatcaggatattcttctaatacctggaatgctgttttcccagggatcgcagtggtgagtaaccatgca

tcatcaggagtacggataaaatgcttgatggtcggaagaggcataaattccgtcagccagtttagtctga

ccatctcatctgtaacatcattggcaacgctacctttgccatgtttcagaaacaactctggcgcatcggg

cttcccatacaatcgatagattgtcgcacctgattgcccgacattatcgcgagcccatttatacccatat

aaatcagcatccatgttggaatttaatcgcggcctagagcaagacgtttcccgttgaatatggctcatac

tcttcctttttcaatattattgaagcatttatcagggttattgtctcatgagcggatacatatttgaatg

tatttagaaaaataaacaaataggggttccgcgcacatttccccgaaaagtgccacctgacgtctaagaa

accattattatcatgacattaacctataaaaataggcgtatcacgaggccctttcgtctcgcgcgtttcg

gtgatgacggtgaaaacctctgacacatgcagctcccggagacggtcacagcttgtctgtaagcggatgc

cgggagcagacaagcccgtcagggcgcgtcagcgggtgttggcgggtgtcggggctggcttaactatgcg

gcatcagagcagattgtactgagagtgcaccatatgcggtgtgaaataccgcacagatgcgtaaggagaa

aataccgcatcagg</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="17">

<INSDSeq>

<INSDSeq_length>43</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..43</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q34">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>agacgctagactagcatgcgactcttcgcgatgtacgggccag</INSDSeq

_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="18">

<INSDSeq>

<INSDSeq_length>43</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..43</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q36">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>caattgttgttgttttgctgtccataaaaccgcccagtagaag</INSDSeq

_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="19">

<INSDSeq>

<INSDSeq_length>44</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..44</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q38">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>tatggacagcaaaacaacaacaattggatctgcgctgattttgt</INSDSe

q_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="20">

<INSDSeq>

<INSDSeq_length>30</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..30</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q40">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>gtcgcatgctagtctagcgtctaaacgcgt</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="21">

<INSDSeq>

<INSDSeq_length>23</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..23</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q42">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>atggacctggaaggcaagcaggg</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="22">

<INSDSeq>

<INSDSeq_length>24</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..24</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q44">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>ccctgcttgccttccaggtccatc</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="23">

<INSDSeq>

<INSDSeq_length>45</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..45</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q46">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>ctggctagccgccaccatggaagacgccaaaaacataaagaaagg</INSDS

eq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="24">

<INSDSeq>

<INSDSeq_length>20</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..20</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q48">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>ggtggcggctagccagcttg</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="25">

<INSDSeq>

<INSDSeq_length>22</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..22</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q50">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>tgtaagggcccgtttaaacccg</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="26">

<INSDSeq>

<INSDSeq_length>38</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..38</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q52">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>cgggtttaaacgggcccttacacggcgatctttccgcc</INSDSeq_sequ

ence>

</INSDSeq>

</SequenceData>

<SequenceData sequenceIDNumber="27">

<INSDSeq>

<INSDSeq_length>5514</INSDSeq_length>

<INSDSeq_moltype>DNA</INSDSeq_moltype>

<INSDSeq_division>PAT</INSDSeq_division>

<INSDSeq_feature-table>

<INSDFeature>

<INSDFeature_key>source</INSDFeature_key>

<INSDFeature_location>1..5514</INSDFeature_location>

<INSDFeature_quals>

<INSDQualifier>

<INSDQualifier_name>mol_type</INSDQualifier_name>

<INSDQualifier_value>other DNA</INSDQualifier_value>

</INSDQualifier>

<INSDQualifier id="q54">

<INSDQualifier_name>organism</INSDQualifier_name>

<INSDQualifier_value>synthetic construct</INSDQualifier_value>

</INSDQualifier>

</INSDFeature_quals>

</INSDFeature>

</INSDSeq_feature-table>

<INSDSeq_sequence>cgcatcaaccaaaccgttattcattcgtgattgcgcctgagcgagacgaaat

acgcgatcgctgttaaaaggacaattacaaacaggaatcgaatgcaaccggcgcaggaacactgccagcg

catcaacaatattttcacctgaatcaggatattcttctaatacctggaatgctgttttcccagggatcgc

agtggtgagtaaccatgcatcatcaggagtacggataaaatgcttgatggtcggaagaggcataaattcc

gtcagccagtttagtctgaccatctcatctgtaacatcattggcaacgctacctttgccatgtttcagaa

acaactctggcgcatcgggcttcccatacaatcgatagattgtcgcacctgattgcccgacattatcgcg

agcccatttatacccatataaatcagcatccatgttggaatttaatcgcggcctagagcaagacgtttcc

cgttgaatatggctcatactcttcctttttcaatattattgaagcatttatcagggttattgtctcatga

gcggatacatatttgaatgtatttagaaaaataaacaaataggggttccgcgcacatttccccgaaaagt

gccacctgacgtctaagaaaccattattatcatgacattaacctataaaaataggcgtatcacgaggccc

tttcgtctcgcgcgtttcggtgatgacggtgaaaacctctgacacatgcagctcccggagacggtcacag

cttgtctgtaagcggatgccgggagcagacaagcccgtcagggcgcgtcagcgggtgttggcgggtgtcg

gggctggcttaactatgcggcatcagagcagattgtactgagagtgcaccatatgcggtgtgaaataccg

cacagatgcgtaaggagaaaataccgcatcaggcgcccctgcaggcagctgcgcgctcgctcgctcactg

aggccgcccgggcaaagcccgggcgtcgggcgacctttggtcgcccggcctcagtgagcgagcgagcgcg

cagagagggagtggccaactccatcactaggggttcctgcggccgcacgcgtttagacgctagactagca

tgcgactcttcgcgatgtacggggccagatatacgcgcgcgttgtatgccaagtacgccccctattgacgt

caatgacggtaaatggcccgcctggcattatgcccagtacatgaccttatgggactttcctacttggcag

tacatctacgtattagtcatcgctattaccatggtgatgcggttttggcagtacatcaatgggcgtggat

agcggtttgactcacggggatttccaagtctccaccccattgacgtcaatgggagtttgttttggcacca

aaatcaacgggactttccaaaatgtcgtaacaactccgccccattgacgcaaatgggcggtaggcgtgta

cggtgggaggtctatataagcagagctggtttagtgaaccgtcagatccgctagtggatccaaagaattc

gggcctccgaaaccatggaagacgccaaaaacataaagaaaggcccggcgccattctatccgctggaaga

tggaaccgctggagagcaactgcataaggctatgaagagatacgccctggttcctggaacaattgctttt

acagatgcacatatcgaggtggacatcacttacgctgagtacttcgaaatgtccgttcggttggcagaag

ctatgaaacgatatgggctgaatacaaatcacagaatcgtcgtatgcagtgaaaactctcttcaattctt

tatgccggtgttgggcgcgttatttatcggagttgcagttgcgcccgcgaacgacatttataatgaacgt

gaattgctcaacagtatgggcatttcgcagcctaccgtggtgttcgtttccaaaaaggggttgcaaaaaa

ttttgaacgtgcaaaaaaagctcccaatcatccaaaaaattattatcatggattctaaaacggattacca

gggatttcagtcgatgtacacgttcgtcacatctcatctacctcccggttttaatgaatacgattttgtg

cccagtccttcgatagggacaagacaattgcactgatcatgaactcctctggatctactggtctgccta

aaggtgtcgctctgcctcatagaactgcctgcgtgagattctcgcatgccagagatcctatttttggcaa

tcaaatcattccggatactgcgattttaagtgttgttccattccatcacggttttggaatgtttactaca

ctcggatatttgatatgtggatttcgagtcgtcttaatgtatagatttgaagaagagctgtttctgagga

gccttcaggattacaagattcaaagtgcgctgctggtgccaaccctattctccttcttcgccaaaagcac

tctgattgacaaatacgatttatctaatttacacgaaattgcttctggtggcgctcccctctctaaggaa

gtcggggaagcggttgccaagaggttccatctgccaggtatcaggcaaggatatgggctcactgagacta

catcagctattctgattacacccgagggggatgataaaccgggcgcggtcggtaaagttgttccatttt

tgaagcgaaggttgtggatctggataccgggaaaacgctgggcgttaatcaaagaggcgaactgtgtgtg

agaggtcctatgattatgtccggttatgtaaacaatccggaagcgaccaacgccttgattgacaaggatg

gatggctacattctggagacatagcttactgggacgaagacgaacacttcttcatcgttgaccgcctgaa

gtctctgattaagtacaaaggctatcaggtggctcccgctgaattggaatccatcttgctccaacacccc

aacatcttcgacgcaggtgtcgcaggtcttcccgacgatgacgccggtgaacttcccgccgccgttgttg

ttttggagcacggaaagacgatgacggaaaaagagatcgtggattacgtcgccagtcaagtaacaaccgc

gaaaaagttgcgcggaggagttgtgtttgtggacgaagtaccgaaaggtcttaccggaaaactcgacgca

agaaaaatcagagagatcctcataaaggccaagaagggcggaaagatcgccgtgtaacccgggtggggta

ccaggtaagtgtacccaattcgccctatagtgagtcgtattacaattcactggccgtcgttttacaacgt

cgtgactgggaaaaccctggcgttacccaacttaatcgccttgcagcacatccccctttcgccagctggc

gtaatagcgaagaggcccgcaccgatcgcccttcccaacagttgcgcagcctgaatggcgaatggagatc

caatttttaagtgtataatgtgttaaactactgattctaattgtttgtgtattttagattcacagtccca

aggctcatttcaggcccctcagtcctcacagtctgttcatgatcataatcagccataccacatttgtaga

ggttttacttgctttaaaaaacctcccacacctccccctgaacctgaaacataaaatgaatgcaattgtt

gttgttaacttgtttattgcagcttataatggttacaaataaagcaatagcatcacaaatttcacaaata

aagcattttttcactgcattctagttgtggtttgtccaaactcatcaatgtatcttaactggcttctac

tgggcggttttatggacagcaagaacaacaacaattggatctgcgctgattttgtaggtaaccacgtgcg

gaccgagcggccgcaggaacccctagtgatggagttggccactccctctctgcgcgctcgctcgctcact

gaggccgggcgaccaaaggtcgcccgacgcccggggctttgcccgggcggcctcagtgagcgagcgagcgc

gcagctgcctgcaggcttggatcccaatggcgcgccgagcttggctcgagcatggtcatagctgtttcct

gtgtgaaattgttatccgctcacaattccacacaacatacgagccggaagcataaagtgtaaagcctggg

gtgcctaatgagtgagctaactcacattaattgcgttgcgctcactgcccgctttccagtcgggaaacct

gtcgtgccagctgcattaatgaatcggccaacgcgcggggagaggcggtttgcgtattgggcgctcttcc

gcttcctcgctcactgactcgctgcgctcggtcgttcggctgcggcgagcggtatcagctcactcaaagg

cggtaatacggttatccacagaatcaggggataacgcaggaaagaacatgtgagcaaaaggccagcaaaa

ggccaggaaccgtaaaaaggccgcgttgctggcgtttttccataggctccgcccccctgacgagcatcac

aaaaatcgacgctcaagtcagaggtggcgaaacccgacaggactataaagataccaggcgtttccccctg

gaagctccctcgtgcgctctcctgttccgaccctgccgcttaccggatacctgtccgcctttctcccttc

gggaagcgtggcgctttctcatagctcacgctgtaggtatctcagttcggtgtaggtcgttcgctccaag

ctgggctgtgtgcacgaaccccccgttcagcccgaccgctgcgccttatccggtaactatcgtcttgagt

ccaacccggtaagacacgacttatcgccactggcagcagccactggtaacaggattagcagagcgaggta

tgtaggcggtgctacagagttcttgaagtggtggcctaactacggctacactagaagaacagtatttggt

atctgcgctctgctgaagccagttaccttcggaaaaagagttggtagctcttgatccggcaaacaaacca

ccgctggtagcggtggttttttgtttgcaagcagcagattacgcgcagaaaaaaaggatctcaagaaga

tcctttgatcttttctacggggtctgacgctcagtggaacgaaaactcacgttaagggattttggtcatg

agattatcaaaaaggatcttcacctagatccttttaaattaaaaatgaagttttaaatcaatctaaagta

tatatgagtaaacttggtctgacagttagaaaaactcatcgagcatcaaatgaaactgcaatttattcat

atcaggattatcaataccatatttttgaaaaagccgtttctgtaatgaaggagaaaactcaccgaggcag

ttccataggatggcaagatcctggtatcggtctgcgattccgactcgtccaacatcaatacaacctatta

atttcccctcgtcaaaaataaggttatcaagtgagaaatcaccatgagtgacgactgaatccggtgagaa

tggcaaaagtttatgcatttctttccagacttgttcaacaggccagccattacgctcgtcatcaaaatca

ct</INSDSeq_sequence>

</INSDSeq>

</SequenceData>

</ST26SequenceListing>

<---

Claims (10)

1. A genetic construct for expression in human or animal cells of a polynucleotide encoding the full-length spike protein of the SARS-CoV-2 virus containing mutations: partial deletion of the RRAR furin site with the removal of all arginines (R) included in the site, as well as mutations K986P, V987P and D614G, characterized by the fact that it is represented by the adeno-associated virus AAV9, into which the said polynucleotide is cloned. 2. The genetic construct according to claim 1, wherein the full-length spike protein of the SARS-CoV-2 virus includes additional mutations inherent to a specific variant of the virus. 3. The genetic construct according to claim 1, wherein the SARS-CoV-2 virus variant is an alpha variant, a beta variant, a delta variant, or an omicron variant. 4. The genetic construct according to claim 1, wherein the full-length spike protein of the SARS-CoV-2 virus has the sequence SEQ ID NO: 2, or SEQ ID NO: 3, or SEQ ID NO: 6. 5. The genetic construct according to claim 1, wherein the polynucleotide encoding the full-length spike protein of the SARS-CoV-2 virus is represented by the sequence SEQ ID NO: 7, or SEQ ID NO: 8, or SEQ ID NO: 11. 6. A recombinant adeno-associated viral vector AAV9 that provides expression in human or animal cells of a polynucleotide encoding the full-length spike protein of the SARS-CoV-2 virus, having a genetic construct according to any of paragraphs 1-5. 7. The recombinant adeno-associated viral vector AAV9 according to claim 6, represented by the sequence SEQ ID NO: 12, or SEQ ID NO: 13, or SEQ ID NO: 16. 8. Use of a genetic construct according to any of claims 1-5 or a recombinant adeno-associated viral vector AAV9 according to any of claims 6, 7 for inducing an immune response against the SARS-CoV-2 virus. 9. Use according to paragraph 8 by intramuscular administration.