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WO2019195975A1 - Procédé de construction pour banque de gènes et son utilisation - Google Patents

Procédé de construction pour banque de gènes et son utilisation Download PDF

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Publication number
WO2019195975A1
WO2019195975A1 PCT/CN2018/082282 CN2018082282W WO2019195975A1 WO 2019195975 A1 WO2019195975 A1 WO 2019195975A1 CN 2018082282 W CN2018082282 W CN 2018082282W WO 2019195975 A1 WO2019195975 A1 WO 2019195975A1
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Prior art keywords
chromosome
biological sample
depth
coverage
heparinase
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English (en)
Chinese (zh)
Inventor
赵佳
杨林
卢森
蒲丹丹
高健
高雅
陈芳
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BGI Shenzhen Co Ltd
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BGI Shenzhen Co Ltd
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Priority to CN201880091404.2A priority Critical patent/CN111868254A/zh
Priority to PCT/CN2018/082282 priority patent/WO2019195975A1/fr
Publication of WO2019195975A1 publication Critical patent/WO2019195975A1/fr
Anticipated expiration legal-status Critical
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C—CHEMISTRY; METALLURGY
    • C40—COMBINATORIAL TECHNOLOGY
    • C40B—COMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES
    • C40B40/00—Libraries per se, e.g. arrays, mixtures
    • C40B40/04—Libraries containing only organic compounds
    • C40B40/06—Libraries containing nucleotides or polynucleotides, or derivatives thereof
    • C40B40/08—Libraries containing RNA or DNA which encodes proteins, e.g. gene libraries
    • C—CHEMISTRY; METALLURGY
    • C40—COMBINATORIAL TECHNOLOGY
    • C40B—COMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES
    • C40B50/00—Methods of creating libraries, e.g. combinatorial synthesis
    • C40B50/06—Biochemical methods, e.g. using enzymes or whole viable microorganisms

Definitions

  • the invention relates to the field of medical detection, in particular to a method for constructing a gene library and an application thereof, in particular to a method for constructing a gene library, a gene library constructed according to the method, and the application in the field of non-invasive prenatal detection of pregnant women .
  • Heparin is a standard method for preventing thrombosis in pregnant women. It has been used clinically to prevent or treat thrombosis in pregnant women for more than 20 years. Heparin therapy can be used to prevent or treat thrombosis and eclampsia.
  • NIPT non-invasive prenatal genetic testing
  • the present invention aims to solve at least to some extent one of the technical problems in the related art, that is, to improve the success rate of non-invasive prenatal genetic testing (NIPT) of a heparin-treated pregnant woman, thereby solving the problem of liver cancer treated by heparin-treated pregnant women.
  • NIPT non-invasive prenatal genetic testing
  • the problem that more heparin-treated pregnant women directly extract and test according to the existing NIPT process, there will be failure to build a database, the results are abnormal, resulting in high blood draw rate, high detection failure rate, which seriously affects NIPT in this category.
  • Application in the crowd It is an object of the present invention to provide a method for constructing a gene library and for use in the field of non-invasive prenatal testing of pregnant women.
  • the present invention provides a method for constructing a gene library suitable for a biological sample containing heparin, comprising the steps of: (1) co-incubating a biological sample with heparinase to obtain A co-incubated sample containing the biological sample and heparinase; (2) obtaining DNA from the co-incubated sample for high-throughput library sequencing, wherein the DNA is amplified by Taq enzyme during the library construction process.
  • a gene library is constructed.
  • the gene library constructed by the method of the invention is especially suitable for the construction of a gene library of a biological sample containing heparin, and is particularly suitable for detecting the abnormality of the fetus in the field of non-invasive prenatal genetic testing for pregnant women.
  • the biological sample in the present invention may be a plasma sample containing heparin.
  • the method of constructing the gene library may further add the following technical features:
  • the heparinase in step (2) is heparinase II.
  • Heparinase II can greatly degrade heparin, thereby minimizing the effect of heparin molecules on the construction of subsequent gene libraries.
  • the heparinase is added in an amount of 0.8 to 2 U per 100 ⁇ L of the biological sample corresponding to the heparinase.
  • the action effect of heparinase can be improved.
  • the heparinase incubation time in step (1) is from 20 to 60 minutes. Thereby, the effect of heparinase can be improved.
  • the reaction system for performing the amplification in the step (2) comprises:
  • the buffer includes dNTP, magnesium ion, Tris-HCl, EDTA-K+.
  • the amplified reaction system may further include other stabilizers for stabilizing the Taq enzyme, such as potassium chloride and the like.
  • the Taq enzyme is Invitrogen Taq enzyme.
  • Invitrogen Taq enzyme is a high-fidelity DNA polymerase, which is a recombinant Taq DNA polymerase, which is composed of Pyrococcus species GB-D polymerase and A mixture of Taq antibodies that is six times more fidelity than Taq DNA polymerase alone and can therefore be used to amplify a range of simple or complex DNA templates.
  • Optimized buffers are also available to increase the fidelity of the enzyme and the amplification efficiency of complex templates.
  • the biological sample is a plasma sample containing heparin.
  • free DNA is extracted from the co-incubated sample to obtain DNA.
  • whole genome DNA is extracted from the co-incubated sample, thereby obtaining DNA.
  • the present invention provides a gene library suitable for use in a blood sample containing heparin, which is constructed according to the method of constructing the gene library of the first aspect of the present invention.
  • a kit comprising a Taq enzyme and heparinase, the kit being adapted for performing an amplification reaction on a biological sample containing heparin.
  • the Taq enzyme and heparinase can be prepared in the form of a kit, which is convenient for those skilled in the art to use.
  • Taq enzymes and heparinase can be packaged separately and can be packaged in individual vials or vials.
  • the amount of Taq enzyme or heparinase contained in each vial or each small tube can be changed according to actual needs, and those skilled in the art can also use the kit to perform amplification reaction on a biological sample containing heparin.
  • the Taq enzyme or heparinase is dispensed or taken out according to actual needs for reaction.
  • the Taq enzyme or heparinase in the kit may be provided in the form of a powder or a liquid preparation according to actual needs.
  • the kit may further comprise a buffer, ddH 2 O, which may comprise dNTP, magnesium ion, Tris-HCl, EDTA-K + . Further, the kit may further include other stabilizers such as potassium chloride and the like which can be used to stabilize the Taq enzyme.
  • a buffer ddH 2 O, which may comprise dNTP, magnesium ion, Tris-HCl, EDTA-K + .
  • the kit may further include other stabilizers such as potassium chloride and the like which can be used to stabilize the Taq enzyme.
  • the above kit may further add the following technical features:
  • a gene library can be constructed according to the method of constructing a gene library according to the first aspect of the present invention using the kit.
  • the kit can be used for non-invasive prenatal genetic testing of pregnant women.
  • the present invention provides a use of a gene library or kit in the field of non-invasive prenatal detection of pregnant women, wherein the gene library is a gene library according to the second aspect of the present invention, the reagent
  • the cassette is a kit according to the third aspect of the invention, preferably in the field of diagnosing fetal genetic abnormalities.
  • the present invention provides a method of constructing a gene library for non-invasive prenatal detection of pregnant women.
  • the method for constructing a gene library for non-invasive prenatal detection of a pregnant woman comprises: obtaining peripheral blood from a pregnant woman, isolating and obtaining plasma as a biological sample, and then constructing a gene according to the method described in the above examples. library.
  • the effect of a certain concentration of heparin on DNA extraction can be effectively eliminated, and the yield and success rate can be increased.
  • Invitrogen TM is replaced during the construction process
  • Taq enzyme can effectively eliminate the inhibitory effect of heparin on DNA polymerase, thereby increasing the success rate of PCR and improving the success rate and quality of database construction.
  • the present invention provides a method for determining a genetic abnormality of a fetus comprising: a) constructing a gene library for obtaining a biological sample using the method of the first aspect of the present invention, respectively, thereby obtaining the biological substance, respectively Sequence information of a plurality of polynucleotide fragments of the sample; b) assigning the plurality of polynucleotide fragments of the biological sample to the chromosome based on the sequence information; c) calculating a depth of coverage of the chromosome based on the sequence information GC content; d) calculating the fitted coverage depth of the chromosome using the GC content of the chromosome and the established relationship between the depth of coverage of the chromosome and the GC content; e) overlaying the fit The depth is compared to the depth of coverage of the chromosomes, wherein differences between them are used to indicate fetal genetic abnormalities.
  • the above method for determining fetal genetic abnormality may further add the following technical features:
  • step d) is the following formula:
  • f(GC i,j ) represents a function of the relationship between the biological sample i, the depth of coverage of chromosome j and the corresponding GC content
  • ⁇ i,j represents the residual of sample i, chromosome j
  • the invention provides a system for determining a genetic abnormality of a fetus comprising:
  • a gene library building unit that constructs a gene library for obtaining a biological sample by using the method described in the first aspect of the present invention, thereby obtaining sequence information of a plurality of polynucleotide fragments of the biological sample, respectively;
  • a computer readable medium for performing the plurality of instructions of the following steps comprising:
  • the fit coverage depth is compared to the coverage depth of the chromosomes, wherein differences between them are used to indicate fetal genetic abnormalities.
  • the above system for determining fetal genetic abnormality may further add the following technical features:
  • the relationship between the depth of coverage of the chromosome and the GC content is the following formula:
  • f(GC i,j ) represents a function of the relationship between the biological sample i, the depth of coverage of chromosome j and the corresponding GC content
  • ⁇ i,j represents the residual of sample i, chromosome j
  • the gene library is constructed by the method of the invention, and is especially suitable for biological samples containing heparin, which can reduce the influence of heparin on gene building.
  • This method can be applied to the field of non-invasive prenatal testing of pregnant women, which can increase the success rate of NIPT detection in pregnant women with heparin treatment, and greatly reduce the phenomenon of heavy blood draw, high GC content and multiple chromosomes.
  • Figure 1 is a graph showing the results of a quality test of the size of a library fragment constructed in an original experiment according to an embodiment of the present invention.
  • Figure 2 is a graph showing the results of a quality test of the size of a library fragment constructed in Experiment 1 according to an embodiment of the present invention.
  • Figure 3 is a graph showing the results of a quality test of the size of a library fragment constructed in Experiment 2 according to an embodiment of the present invention.
  • Figure 4 is a graph showing the results of a quality test of the size of a library fragment constructed in Experiment 3 according to an embodiment of the present invention.
  • the present invention provides a method for constructing a gene library suitable for a biological sample containing heparin, thereby reducing the influence of heparin molecules on gene building, thereby improving the accuracy and accuracy of gene detection.
  • the method mainly reduces the heparin molecule by adding heparinase treatment to a biological sample containing heparin; and simultaneously using Taq enzyme for DNA polymerization, thereby increasing the efficiency of amplification and reducing the influence of heparin on the polymerase during DNA polymerization.
  • the present invention provides a method for constructing a gene library suitable for a sample containing heparin, comprising the following steps: (1) co-incubating a biological sample with heparinase to obtain a content Co-incubation samples of biological samples and heparinase; (2) obtaining DNA from the co-incubated samples for high-throughput library sequencing, wherein the DNA is amplified by Taq enzyme during database construction.
  • the method of the invention is especially suitable for the field of non-invasive prenatal testing of pregnant women. It should be noted that the construction of the gene library of the present invention is not limited to plasma free DNA, and can also be used for the extraction of whole genome DNA in a heparin-treated population.
  • the treatment of whole blood samples with heparinase is also effective, and inhibits heparin to DNA. Extraction and the effects of PCR.
  • the DNA comprises both maternal DNA and fetal DNA when tested against a pregnant woman.
  • the invention is applicable to all PCR reactions with heparin molecules, and the Taq enzyme can effectively prevent the inhibition of heparin on DNA polymerase during the amplification process.
  • heparinase can be added to the biological sample to obtain a co-incubated sample containing the biological sample and heparinase.
  • a biological sample can also be added to heparinase to obtain a co-incubated sample containing the biological sample and heparinase.
  • heparinase can be dispensed into small vials and the biological sample can be added directly to the vial.
  • Taq enzyme is a thermostable DNA polymerase in Thermus aquaticus, which can withstand high temperatures above 90 °C without inactivation, which greatly reduces the cost of PCR technology.
  • the Taq enzyme can also be referred to as Taq DNA polymerase.
  • the obtained DNA fragments can be sequenced using second-generation sequencing and three-generation sequencing techniques.
  • the sequencing described in the present invention can be applied to a sequencing sequencing platform such as a Roche 454GS FLX sequencing platform, a Solexa Genome sequencing platform, and an Ion Torrent sequencing platform, and can also be applied to a connection sequencing platform such as an Analyzer sequencing platform and an ABI SOLID sequencing platform.
  • the present invention provides a method for constructing a gene library for non-invasive prenatal detection of pregnant women, comprising:
  • peripheral blood from a pregnant woman, separating plasma to obtain a biological sample, adding heparinase to the biological sample, and (2) extracting free DNA in the biological sample, obtaining a DNA fragment, and performing high-throughput
  • the library was sequenced, and the DNA fragment was amplified by Taq enzyme during the database construction process.
  • the content of fetal free DNA (cffDNA) in the peripheral blood of pregnant women accounts for about 5-10% of the total free DNA, and is about 150 bp long. It can be detected in 4 weeks of pregnancy.
  • Peripheral blood is obtained from pregnant women to obtain fetal free DNA nucleic acid information in the peripheral blood.
  • the inventors of the present invention have found that blood samples of these people can be added to heparinase treatment before the establishment of the library, thereby degrading heparin and reducing the influence of heparin on the construction of the library; Amplification of the target DNA fragment can improve the efficiency of amplification, and thus can be used to solve the failure of heparin-containing blood sample construction.
  • Gene libraries constructed using this method are used for non-invasive prenatal genetic testing in pregnant women, and in particular to determine fetal genetic abnormalities. For example, it can be used to detect or determine whether the fetus has an abnormal number of chromosomes in the presence of aneuploidy syndrome.
  • the non-invasive prenatal test in the present invention refers to a technique for detecting the fetus before giving birth without causing any trauma to the fetus.
  • the number of chromosomes of aneuploidy syndrome in the present invention is abnormal, including but not limited to 21-trisomy, 18-trisomy, 13-trisomy, and X chromosome number abnormalities.
  • Aneuploidy refers to an abnormal number of chromosomes that increase or decrease the entire chromosome of a part of a genome. Unlike chromosomal euploidy changes that lead to miscarriage and structural changes in some chromosomes, aneuploidy abnormalities in some chromosomes can produce viable birth defects.
  • a system for determining fetal genetic abnormalities, or a computer scale medium for determining fetal genetic abnormalities, or for determining a fetus may be used.
  • the method of genetic abnormality is used for diagnosis. This method can remove the defect of GC deviation of sequencing result due to the difference of GC content of chromosome, and make the diagnosis result more accurate.
  • part or all of the contents of the patent application No. 201180067286.X, the authorization number being CN103403183B can be cited in the present invention.
  • a person skilled in the art may select part or all of the contents of the patent (application number 201180067286.X, authorization number CN103403183B) according to actual needs.
  • whether the fetus is genetically abnormal can be determined according to a computer readable medium or system for diagnosing fetal genetic abnormalities as described in Patent No. CN103403183B.
  • Methods for diagnosing fetal genetic abnormalities using the computer readable medium or system include:
  • step d) is the following formula:
  • f(GC i,j ) represents a function of the relationship between the biological sample i, the depth of coverage of chromosome j and the corresponding GC content
  • ⁇ i,j represents the residual of sample i, chromosome j
  • the method can be used for detecting fetal chromosomal abnormalities, and is particularly suitable for detecting aneuploidy, polyploidy, monosomy, trisomy, trisomy 21, trisomy, 14 trisomy, 15 three Body, 16 trisomy, 18 trisomy, 22 trisomy, triploid, tetraploid, and sex chromosome abnormalities include XO, XXY, XYY, and XXX.
  • Specific regions in the human genome can also be focused on according to the method to identify partial and partial trisomies.
  • the method can involve analyzing sequence data in a determined chromosome sliding "window", such as a continuous, non-overlapping 50 kb region distributed over the entire chromosome.
  • Partial trisomy 13q, 8p (8p23.1), 7q, distal 6p, 5p, 3q (3q25.1), 2q, lq (lq42.1 and lq21-qter), part have been reported, among others Xpand monomeric 4q35.1.
  • 18q21.1-qter repeats partial duplication of the long arm of chromosome 18 can lead to Edwards syndrome (Mewar, et al., Am J Hum Genet. (1993) 53: 1269-78).
  • the fetal fraction is estimated based on the sequence information obtained from the polynucleotide fragment of the sample.
  • the depth of coverage and GC content of the X and Y chromosomes can be used to estimate the fetal fraction.
  • the fetal gender is determined based on the sequence information obtained from the polynucleotide fragment of the sample.
  • the depth of coverage and GC content of the X and Y chromosomes can be used to determine the sex of the fetus.
  • the comparison of the fit coverage depth to the coverage depth of the chromosome is performed by a statistical hypothesis test, wherein one hypothesis is that the fetus is euploid (H0) and the other It is assumed that the fetus is aneuploid (H1).
  • the student t-statistics are calculated for the two hypotheses, respectively, as tl and t2.
  • the log likelihood ratios of t1 and t2 are calculated.
  • a log likelihood ratio > 1 indicates that the fetal trisomy is indicated.
  • Plasma separation using EDTA anticoagulation tube, take 5mL pregnant women's peripheral blood according to the peripheral blood standard collection operation, please slowly invert 10 times after blood collection to mix the blood and the tube components, and centrifuge at 1600g for 10 minutes at 4 °C. The supernatant was dispensed onto a plurality of 2.0 mL centrifuge tubes on an ice box. After centrifugation at 16000 g for 10 minutes at 4 ° C, the resulting supernatant was transferred to a new numbered 2.0 mL centrifuge tube on an ice box, and each centrifuge tube was transferred to 600 ⁇ L of plasma.
  • the EDTA anticoagulant tube refers to a tube containing ethylenediaminetetraacetic acid (EDTA) and/or a salt thereof, and the ethylenediaminetetraacetic acid and the salt thereof are an amino acid polycarboxylic acid, which can effectively chelate the blood sample.
  • EDTA ethylenediaminetetraacetic acid
  • the salt thereof are an amino acid polycarboxylic acid, which can effectively chelate the blood sample.
  • Calcium ions, chelation of calcium or removal of the calcium reaction site will block and terminate the endogenous or exogenous coagulation process, thereby preventing the blood sample from solidifying.
  • Plasma free DNA was extracted, and DNA was extracted according to the Magen MagPure Circulating DNA Mini KF Kit (Cat. No. MD5427-02).
  • the extracted DNA was constructed using the fetal chromosome aneuploidy (T21, T18, T13) detection kit (BGI MFG, article number: MFG030008).
  • a high-fidelity hot-start polymerase premix (KAPA HiFi HotStart ReadyMix) is amplified for the sequencing library during DNA amplification.
  • the premix includes KAPA HiFi HotStart DNase (0.5 U/25 ⁇ L), dNTPs (0.3 mM), MgCl 2 (2.5 mM), and a buffer that stabilizes the KAPA HiFi HotStart DNA
  • the action of the enzyme includes 100 mM Tris-HCl (pH 8.3) and 500 mM potassium chloride.
  • the constructed gene library was then sequenced using the BGISEQ-500 High Throughput Sequencing Kit (SE50) V2.1, Box 3 (RT) (BGI MGI, Cat. No. 85-05239-00).
  • step 4 The sequencing data in step 4 is analyzed according to the method described in the patent CN103403183B, and the corresponding result is obtained, thereby detecting the aneuploidy of the chromosome.
  • the sample information is shown in Table 1.
  • the reasons for the failure of the test include: failure to build a library, low library concentration; high GC content, QC does not pass; or low fetal concentration, or multiple chromosomes out of range, unable to analyze the three-body signal.
  • the inventors have improved the library construction method in the first embodiment.
  • the improved method is used to rebuild the library, and the following experiments are respectively designed, as follows:
  • Improved experiment 1 extracting DNA after adding heparinase II to the extracted maternal plasma
  • step a is added between step 1 and step 2 of the original experiment.
  • Step a In 600 ⁇ L of heparin-treated pregnant women, 7.4 U of heparinase II (Sigma) was added and incubated at 37 ° C for 30 min.
  • the reaction system is as follows: 20 ⁇ L DNA, 5 ⁇ L 10 ⁇ PCR buffer, 2.5 ⁇ L MgCl 2 , 2.5 ⁇ L dNTP (100 ⁇ mol), 4 ⁇ L PCR primer, 0.5 ⁇ L Taq, 15.5 ⁇ L ddH 2 O.
  • invitrogen TM Taq enzyme included Taq high fidelity DNA polymerase, 20 mM Tris-HCl (pH 8.0), 0.1 mM EDTA, 1 mM DTT and 50% (v/v) glycerol and used to stabilize Taq Stabilizer for high fidelity DNA polymerase.
  • step a is added between step 1 and step 2 of the original experiment.
  • Step a In 600 ⁇ L of heparin-treated pregnant women, 7.4 U of heparinase II (Sigma) was added and incubated at 37 ° C for 30 min; the heparin molecules remaining in the plasma were digested.
  • Example 1 The libraries constructed in Example 1 and Example 2 were tested separately.
  • the quality of the library was >3ng/ ⁇ L, and the library was qualified. See Table 2 for details.
  • KAPA HiFi HotStart DNA polymerase in the high-fidelity hot start polymerase master mix has a higher concentration of the DNA library constructed using the Taq enzyme.
  • KAPA HiFi HotStart DNA Polymerase caused a part of DNA loss during the process of improving fidelity, so that the concentration of the constructed DNA library was not high.
  • This situation is extremely unsuitable for non-invasive prenatal testing of pregnant women. Because the non-invasive prenatal testing of pregnant women, because of the limited DNA information about the fetus given in the DNA information of pregnant women, it is necessary to obtain a DNA library of the most comprehensive and higher amount possible in order to obtain more reliable data. Of course, this result can also be seen from the analysis of the sequencing results in the above step 3.
  • the DNA libraries constructed in the first embodiment and the second embodiment are respectively sequenced, and the plurality of multinuclei of the biological sample is firstly determined based on the sequence information by using the method described in the patent CN103403183B.
  • the nucleotide fragment is assigned to the chromosome; the coverage depth and GC content of the chromosome are calculated based on the sequence information; and the relationship between the GC content of the chromosome and the established depth of coverage and GC content of the chromosome is used to calculate
  • the fit of the chromosome covers the depth; the fit coverage depth is compared to the depth of coverage of the chromosome, wherein differences between them are used to indicate fetal genetic abnormalities.
  • the statistic Z value of a certain chromosome is calculated, wherein the Z value ⁇ 3 is used as a threshold for diagnosing a chromosome of a sample to be tested, indicating 99% credible three-body abnormality, Z value ⁇ -3, as a diagnosis
  • the threshold of a chromosome monomer in the sample is measured, indicating that 99% of the authentic monomer is abnormal.
  • the fetal fraction in each sample is also calculated to characterize the concentration of the fetus in the sample. The higher the fetal fraction, the more reliable the results obtained.
  • Sample 1 0.4398 0.051 1.438 -2.717 -0.675
  • Sample 2 0.4357 0.023 1.454 1.037 -0.337
  • Sample 3 0.4433 0.0515 0.668 -1.277 1.96
  • Sample 4 0.4268 0.0436 -1.762 1.91 -0.474
  • Sample 5 0.4209 0.034 -0.923 0.879 1.444
  • Sample 6 0.4284 0.056 1.96 -0.719 -0.07
  • Sample 1 0.4132 0.055 0.897 -1.43 -0.903
  • Sample 2 0.4366 0.043 1.943 1.011 0.486
  • Sample 3 0.4428 0.052 -0.983 0.453 0.345
  • Sample 4 0.4321 0.0563 -1.894 0.47 1.95
  • Sample 5 0.4105 0.065 -0.768 0.635 -0.345
  • Sample 6 0.4254 0.068 1.23 1.34 0.21

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Abstract

L'invention concerne un procédé de construction d'une bibliothèque de gènes, une banque de gènes construite selon le procédé, et une utilisation dans le domaine des tests prénataux non invasifs de femmes enceintes. Le procédé de construction d'une banque de gènes comprend : la co-incubation d'un échantillon biologique et d'héparinase pour obtenir un échantillon de co-incubation comprenant l'échantillon biologique et l'héparinase; et l'acquisition d'ADN à partir de l'échantillon de co-incubation et la mise en oeuvre d'un séquençage et d'une construction de banque à haut rendement, l'amplification d'ADN étant effectuée à l'aide d'une enzyme Taq dans le processus de construction de banque.
PCT/CN2018/082282 2018-04-09 2018-04-09 Procédé de construction pour banque de gènes et son utilisation Ceased WO2019195975A1 (fr)

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CN201880091404.2A CN111868254A (zh) 2018-04-09 2018-04-09 基因文库的构建方法及其应用
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CN112522387B (zh) * 2020-12-10 2022-05-20 北京优迅医学检验实验室有限公司 一种无创产前检测染色体异常的装置

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