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WO2019074461A2 - Development of molecular kit based on kasp system for tm-2a gene which provides resistance against tomato mosaic virus - Google Patents

Development of molecular kit based on kasp system for tm-2a gene which provides resistance against tomato mosaic virus Download PDF

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Publication number
WO2019074461A2
WO2019074461A2 PCT/TR2018/050531 TR2018050531W WO2019074461A2 WO 2019074461 A2 WO2019074461 A2 WO 2019074461A2 TR 2018050531 W TR2018050531 W TR 2018050531W WO 2019074461 A2 WO2019074461 A2 WO 2019074461A2
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WIPO (PCT)
Prior art keywords
gene
kasp
mosaic virus
resistance against
marker
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/TR2018/050531
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French (fr)
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WO2019074461A3 (en
Inventor
Zubeyir DEVRAN
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
MY Genetik Tarim Teknoloji Lab Tic Ltd Sti
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MY Genetik Tarim Teknoloji Lab Tic Ltd Sti
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Publication of WO2019074461A2 publication Critical patent/WO2019074461A2/en
Publication of WO2019074461A3 publication Critical patent/WO2019074461A3/en
Anticipated expiration legal-status Critical
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    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12QMEASURING 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
    • C12Q1/00Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6876Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6888Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms
    • C12Q1/6895Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms for plants, fungi or algae
    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12QMEASURING 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
    • C12Q2600/00Oligonucleotides characterized by their use
    • C12Q2600/13Plant traits
    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12QMEASURING 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
    • C12Q2600/00Oligonucleotides characterized by their use
    • C12Q2600/156Polymorphic or mutational markers

Definitions

  • the present invention relates to a marker developed for Tm2a gene which provides resistance against tomato mosaic virus and relates to the kits comprising said marker and relates to the usage of said marker in the detection of the presence of Tm-2 2 (Tm-2 a ) gene which provides resistance against tomato mosaic virus in tomato plant.
  • Tm-2 a Tm-2 2
  • Tomato is one of the most important plants essentially grown in the world. Various diseases and harmful insects lead to economic losses in tomato. Tomato mosaic virus (ToMV) is one of the most important of these diseases. The most successful method against this disease is to use resistant kinds. As known, resistance in plants is provided by R genes. In tomato plants, the resistance against tomato mosaic virus is provided by the Tm-2 2 gene. The detection of whether there is resistance gene in plants or not can be determined by means of inoculation of plants with pathogen or by means of molecular markers. The inoculation studies of plants with pathogen take time and need dense labor and a climate-controlled feeding cabinet is needed. Molecular markers can be used as a method which is alternative to inoculation with pathogen.
  • Said markers can be primers which are developed by the own sequencing of the gene or said markers can be developed from a DNA region which is substantially close to the gene.
  • These DNA markers can be analyzed by means of PCR method.
  • errors may occur in reading the DNA bands and dense labor and time are needed. As a result, because of all of the abovementioned problems, an improvement is required in the related technical field.
  • the present invention relates to markers developed by using KASP system, for eliminating the above mentioned disadvantages and for bringing new advantages to the related technical field.
  • An object of the present invention is to provide a rapid, repeatable, low-cost and reliable molecular marker.
  • the present invention relates to Tm-2 2 gene KASP marker which provides resistance against tomato mosaic virus.
  • the present invention gives successful results in the marker adjuvant selection of Tm-2 2 gene in tomato improvement.
  • Tm-2 2 gene KASP marker which provides resistance against tomato mosaic virus
  • the subject matter Tm-2 2 gene KASP marker has the DNA array of GAGAAGTGCTCAWTTCTATGAATCTCAACGTTGAGA[T/A]TTTGGTCTTAGTT CAGTATTTTTTTGTATAATTGGAT (SEQ ID NO: 1 ).
  • SNP array of said DNA array is as follows: ATCTCAACGT TGAGA[T/A]TTTG GTCTTAGTTC (SEQ ID NO: 2).
  • Tm-2 2 gene which provides resistance against tomato mosaic virus, can be defined in a rapid and reliable manner.
  • KAPS used within the scope of the present invention is the shortened form of "Kompetitive allele specific PCR” statement and it is a homogeneous and fluorescence- based derivative of polymerase chain reaction. This method is based on allele-based oligo extensions in signal production and fluorescent resonance energy transfer.
  • kits comprising the subject matter KASP markers for the Tm-2 2 gene.
  • DNA array of the KASP markers for the Tm-2 2 gene existing in the subject matter kit has the following form:
  • GAGAAGTGCTCAWTTCTATGAATCTCAACGTTGAGA[T/A]TTTGGTCTTAGTTCAGTA TTTTTTTGT AT A ATTG GAT SEQ ID NO: 1 .
  • SNP array of the KASP markers for the Tm-2 2 gene existing in the subject matter kit has the following form: ATCTCAACGTTGAGA[T/A]TTTGGTCTTAGTTC (SEQ ID NO: 2).
  • auxiliary substances can exist in addition to the abovementioned DNA array and/or SNP array.
  • the present invention relates to a method to be used in obtaining KASP markers for the Tm-2 2 gene which is compliant to the subject matter and said method comprises the following steps:
  • the present invention relates to the usage of KASP marker, which has the DNA array of
  • the present invention moreover relates to the usage of KASP marker, which has the SNP array of ATCTCAACGTTGAGA[T/A]TTTGGTCTTAGTTC (SEQ ID NO: 2), in detection of Tm-2 2 gene in tomato plant.

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  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Engineering & Computer Science (AREA)
  • Organic Chemistry (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Health & Medical Sciences (AREA)
  • Biotechnology (AREA)
  • Zoology (AREA)
  • Wood Science & Technology (AREA)
  • Immunology (AREA)
  • Mycology (AREA)
  • Microbiology (AREA)
  • Molecular Biology (AREA)
  • Botany (AREA)
  • Biophysics (AREA)
  • Physics & Mathematics (AREA)
  • Biochemistry (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

The present invention relates to a marker developed for Tm-22 (Tm2a) gene which provides resistance against tomato mosaic virus and relates to the kits comprising said marker and relates to the usage of said marker in the detection of the presence of Tm-22 gene which provides resistance against tomato mosaic virus in tomato plant.

Description

DEVELOPMENT OF MOLECULAR KIT BASED ON KASP SYSTEM FOR TM-2A GENE WHICH PROVIDES RESISTANCE AGAINST TOMATO MOSAIC VIRUS
TECHNICAL FIELD
The present invention relates to a marker developed for Tm2a gene which provides resistance against tomato mosaic virus and relates to the kits comprising said marker and relates to the usage of said marker in the detection of the presence of Tm-22 (Tm-2a) gene which provides resistance against tomato mosaic virus in tomato plant.
PRIOR ART Tomato is one of the most important plants essentially grown in the world. Various diseases and harmful insects lead to economic losses in tomato. Tomato mosaic virus (ToMV) is one of the most important of these diseases. The most successful method against this disease is to use resistant kinds. As known, resistance in plants is provided by R genes. In tomato plants, the resistance against tomato mosaic virus is provided by the Tm-22 gene. The detection of whether there is resistance gene in plants or not can be determined by means of inoculation of plants with pathogen or by means of molecular markers. The inoculation studies of plants with pathogen take time and need dense labor and a climate-controlled feeding cabinet is needed. Molecular markers can be used as a method which is alternative to inoculation with pathogen. Said markers can be primers which are developed by the own sequencing of the gene or said markers can be developed from a DNA region which is substantially close to the gene. These DNA markers can be analyzed by means of PCR method. However, after gel electrophoresis process of PCR products, errors may occur in reading the DNA bands and dense labor and time are needed. As a result, because of all of the abovementioned problems, an improvement is required in the related technical field.
BRIEF DESCRIPTION OF THE INVENTION The present invention relates to markers developed by using KASP system, for eliminating the above mentioned disadvantages and for bringing new advantages to the related technical field. An object of the present invention is to provide a rapid, repeatable, low-cost and reliable molecular marker.
In order to realize all of the abovementioned objects and the objects which are to be deducted from the detailed description below, the present invention relates to Tm-22 gene KASP marker which provides resistance against tomato mosaic virus. Thus, the present invention gives successful results in the marker adjuvant selection of Tm-22 gene in tomato improvement. DETAILED DESCRIPTION OF THE INVENTION
In this detailed description, the subject matter Tm-22 gene KASP marker, which provides resistance against tomato mosaic virus, is explained with references to examples without forming any restrictive effect only in order to make the subject more understandable.
In an application of the invention, the subject matter Tm-22 gene KASP marker has the DNA array of GAGAAGTGCTCAWTTCTATGAATCTCAACGTTGAGA[T/A]TTTGGTCTTAGTT CAGTATTTTTTTGTATAATTGGAT (SEQ ID NO: 1 ). In another application of the invention, SNP array of said DNA array is as follows: ATCTCAACGT TGAGA[T/A]TTTG GTCTTAGTTC (SEQ ID NO: 2).
By means of the usage of the subject matter KASP markers, Tm-22 gene, which provides resistance against tomato mosaic virus, can be defined in a rapid and reliable manner.
The "KAPS" statement used within the scope of the present invention is the shortened form of "Kompetitive allele specific PCR" statement and it is a homogeneous and fluorescence- based derivative of polymerase chain reaction. This method is based on allele-based oligo extensions in signal production and fluorescent resonance energy transfer.
The "SNP" statement used within the scope of the present invention is the shortened form of "Single Nucleotide Polymorphism" statement and it is described as each nucleotide difference between two DNA sequences. From another perspective, the present invention relates to kits comprising the subject matter KASP markers for the Tm-22 gene. DNA array of the KASP markers for the Tm-22 gene existing in the subject matter kit has the following form:
GAGAAGTGCTCAWTTCTATGAATCTCAACGTTGAGA[T/A]TTTGGTCTTAGTTCAGTA TTTTTTTGT AT A ATTG GAT (SEQ ID NO: 1 ).
SNP array of the KASP markers for the Tm-22 gene existing in the subject matter kit has the following form: ATCTCAACGTTGAGA[T/A]TTTGGTCTTAGTTC (SEQ ID NO: 2).
In the subject matter kit; solvent, etc. auxiliary substances can exist in addition to the abovementioned DNA array and/or SNP array.
From another perspective, the present invention relates to a method to be used in obtaining KASP markers for the Tm-22 gene which is compliant to the subject matter and said method comprises the following steps:
a) Selecting tomato materials
b) PCR study by means of primers
c) Sequencing analysis of PCR products
d) Determining SNP regions in resistant and sensitive plants
e) Developing molecular markers with KASP technology.
From another perspective, the present invention relates to the usage of KASP marker, which has the DNA array of
GAGAAGTGCTCAWTTCTATGAATCTCAACGTTGAGA[T/A]TTTGGTCTTAGTTC AGTA TTTTTTTGT AT AATTGGAT (SEQ ID NO: 1 ), in detection of Tm-22 gene in tomato plant.
The present invention moreover relates to the usage of KASP marker, which has the SNP array of ATCTCAACGTTGAGA[T/A]TTTGGTCTTAGTTC (SEQ ID NO: 2), in detection of Tm-22 gene in tomato plant.
The protection scope of the present invention is set forth in the annexed claims and cannot be restricted to the illustrative disclosures given above, under the detailed description. It is because a person skilled in the relevant art can obviously produce similar embodiments under the light of the foregoing disclosures, without departing from the main principles of the present invention.

Claims

1. A marker compliant for use in detection of Tm-22 gene which provides resistance against tomato mosaic virus, wherein it is KASP marker which has the DNA array of GAGAAGTGCTCAWTTCTATGAATCTCAACGTTGAGA[T/A]TTTGGTCTTAGTTCAG TA TTTTTTTGTATAATTGGAT (SEQ ID NO: 1 ).
2. A marker according to claim 1 , wherein it has the SNP array of ATCTCAACGTTGAGA[T/A]TTTGGTCTTAGTTC (SEQ ID NO: 2).
3. A kit compliant for use in detection of Tm-22 gene which provides resistance against tomato mosaic virus, wherein it has KASP marker having the DNA array of GAGAAGTGCTCAWTTCTATGAATCTCAACGTTGAGA[T/A]TTTGGTCTTAGTTCAG TA TTTTTTTGTATAATTGGAT (SEQ ID NO: 1 ) therein.
4. A kit according to claim 3, wherein it has KASP marker which has SNP array of ATCTCAACGTTGAGA[T/A]TTTGGTCTTAGTTC (SEQ ID NO: 2) therein.
5. Usage of KASP marker, which has DNA array of GAGAAGTGCTCAWTTCTATGAATCTCAACGTTGAGA[T/A]TTTGGTCTTAGTTCAG TA TTTTTTTGTATAATTGGAT (SEQ ID NO: 1 ), in detection of Tm-22 gene in tomato plant.
6. Usage of KASP marker, which has SNP array of ATCTCAACGTTGAGA[T/A]TTTGGTCTTAGTTC (SEQ ID NO: 2), in detection of Tm- 22 gene in tomato plant.
PCT/TR2018/050531 2017-10-10 2018-09-27 Development of molecular kit based on kasp system for tm-2a gene which provides resistance against tomato mosaic virus Ceased WO2019074461A2 (en)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
TR2017/15272A TR201715272A2 (en) 2017-10-10 2017-10-10 DEVELOPMENT OF A MOLECULAR KIT BASED ON THE CASTING SYSTEM FOR Tm-22 GENI WHICH PROVIDES RESISTANCE AGAINST TOMATO MOSAIC VIRUS IN TOMATO PLANT
TR2017/15272 2017-10-10

Publications (2)

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WO2019074461A2 true WO2019074461A2 (en) 2019-04-18
WO2019074461A3 WO2019074461A3 (en) 2019-05-09

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN113832251A (en) * 2021-09-30 2021-12-24 中国农业科学院农业基因组研究所 Combination of SNP loci for detecting resistance to tomato mosaic virus disease and its application

Family Cites Families (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR101835124B1 (en) * 2015-12-03 2018-03-06 서울대학교산학협력단 KASP primer set based on SNP for discriminating pepper disease-resistant cultivar and uses thereof

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN113832251A (en) * 2021-09-30 2021-12-24 中国农业科学院农业基因组研究所 Combination of SNP loci for detecting resistance to tomato mosaic virus disease and its application
CN113832251B (en) * 2021-09-30 2024-04-19 中国农业科学院农业基因组研究所 SNP locus combination for detecting tomato mosaic virus resistance and application thereof

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TR201715272A2 (en) 2017-10-23

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