AU2021102786A4 - Snp molecular marker site for resistance to cucumber powdery mildews and application thereof - Google Patents
Snp molecular marker site for resistance to cucumber powdery mildews and application thereof Download PDFInfo
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- 241000221785 Erysiphales Species 0.000 title claims abstract description 31
- 235000010799 Cucumis sativus var sativus Nutrition 0.000 title claims abstract description 28
- 239000003147 molecular marker Substances 0.000 title claims abstract description 16
- 244000299906 Cucumis sativus var. sativus Species 0.000 title abstract 2
- -1 pplication Species 0.000 title description 6
- 240000008067 Cucumis sativus Species 0.000 claims abstract description 33
- 238000000034 method Methods 0.000 claims abstract description 9
- 238000012408 PCR amplification Methods 0.000 claims description 8
- 230000003321 amplification Effects 0.000 claims description 5
- 238000003199 nucleic acid amplification method Methods 0.000 claims description 5
- 238000009472 formulation Methods 0.000 claims description 4
- 239000000203 mixture Substances 0.000 claims description 4
- 239000002773 nucleotide Substances 0.000 claims description 4
- 125000003729 nucleotide group Chemical group 0.000 claims description 4
- 208000035240 Disease Resistance Diseases 0.000 claims description 3
- 102000003960 Ligases Human genes 0.000 claims description 3
- 108090000364 Ligases Proteins 0.000 claims description 3
- 239000011535 reaction buffer Substances 0.000 claims description 3
- 235000009849 Cucumis sativus Nutrition 0.000 abstract description 7
- 230000035945 sensitivity Effects 0.000 abstract 1
- 201000010099 disease Diseases 0.000 description 16
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 16
- 239000000463 material Substances 0.000 description 13
- 108090000623 proteins and genes Proteins 0.000 description 8
- 241000196324 Embryophyta Species 0.000 description 7
- 238000011081 inoculation Methods 0.000 description 7
- 238000009395 breeding Methods 0.000 description 3
- 230000001488 breeding effect Effects 0.000 description 3
- 238000005251 capillar electrophoresis Methods 0.000 description 3
- 238000001514 detection method Methods 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- 239000005547 deoxyribonucleotide Substances 0.000 description 2
- 239000000243 solution Substances 0.000 description 2
- 239000007921 spray Substances 0.000 description 2
- 239000004743 Polypropylene Substances 0.000 description 1
- RWQNBRDOKXIBIV-UHFFFAOYSA-N Thymine Natural products CC1=CNC(=O)NC1=O RWQNBRDOKXIBIV-UHFFFAOYSA-N 0.000 description 1
- 208000005652 acute fatty liver of pregnancy Diseases 0.000 description 1
- 238000012271 agricultural production Methods 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 229940104302 cytosine Drugs 0.000 description 1
- OPTASPLRGRRNAP-UHFFFAOYSA-N cytosine Natural products NC=1C=CNC(=O)N=1 OPTASPLRGRRNAP-UHFFFAOYSA-N 0.000 description 1
- 238000001962 electrophoresis Methods 0.000 description 1
- 239000012634 fragment Substances 0.000 description 1
- 238000001502 gel electrophoresis Methods 0.000 description 1
- 230000002068 genetic effect Effects 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000009335 monocropping Methods 0.000 description 1
- 239000013642 negative control Substances 0.000 description 1
- 239000000447 pesticide residue Substances 0.000 description 1
- 229920001155 polypropylene Polymers 0.000 description 1
- 239000013641 positive control Substances 0.000 description 1
- 230000000750 progressive effect Effects 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 229940113082 thymine Drugs 0.000 description 1
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- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
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- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6888—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms
- C12Q1/6895—Nucleic 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
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H1/00—Processes for modifying genotypes ; Plants characterised by associated natural traits
- A01H1/04—Processes of selection involving genotypic or phenotypic markers; Methods of using phenotypic markers for selection
- A01H1/045—Processes of selection involving genotypic or phenotypic markers; Methods of using phenotypic markers for selection using molecular markers
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- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/156—Polymorphic or mutational markers
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Abstract
The present invention discloses an SNP molecular marker for resisting
cucumber powdery mildews and designs an identification primer and a kit
according to an SNP site. Through an identification method of the present
invention, the resistance of cucumbers to powdery mildews may be rapidly
identified; and the identification method is strong in specificity and high in
sensitivity.
Drawings of Description
2 3 4 5 6 7 8 9 10 11 12 13 14 15
Fig. 1
1
Description
Drawings of Description
2 3 4 5 6 7 8 9 10 11 12 13 14 15
Fig. 1
Description
Technical Field
The present invention belongs to the technical field of molecular detection, and particularly relates to an SNP molecular marker site for resisting cucumber powdery mildews and an application thereof.
Background
Cucumber powdery mildew is a widespread worldwide disease as well as one of major foliage diseases of cucumbers. The cucumber powdery mildew is short in latent period and strong in epidemicity, manually occurs and severely influences the yield and the quality of the cucumbers. At present, a main control measure of the cucumber powdery mildews is chemical control. However, a pesticide residue of a product is a non-negligible safety problem, and particularly, the problem of continuous cropping obstacles in cucumber production is more and more serious after cultivation is conducted by using facilities. Thus, selection of disease-resistant varieties is still an effective way of controlling the powdery mildews fundamentally. The advent of a molecular marker technology enables accurate detection of genes and accurate transfer of target genes to come true. Researchers primarily locate anti-disease genes by using molecular marker technologies such as AFLP and SSR. However, a genetic distance between the anti-disease genes and the molecular marker is relatively large generally, which indicates that the resistant molecule mechanism of the cucumber powdery mildews is very complex, and there are more scientific problems to be studied and explored. By fully excavating and utilizing these resistance genes, resistance gene resources in resistance breeding of the cucumber powdery mildews will be enriched, and an important
Description
basis will be provided for introducing excellent resistance genes into resistance breeding of the cucumber powdery mildews. At present, SNP sites of the cucumber powdery mildews discovered by those skilled in the art have certain problems. Thus, how to provide an SNP molecular marker site for resistance to cucumber powdery mildews and am application thereof is a problem needing to be solved urgently in the field.
Summary The present invention discloses an SNP molecular marker site for resistance to cucumber powdery mildews and an application thereof. In order to achieve the above purpose, the present invention adopts the following technical solution: An application of an SNP sequence for resisting the cucumber powdery mildews in preparing a formulation for detecting resistance to the cucumber powdery mildews is provided, wherein position 600 of a nucleotide sequence of an SNP molecular marker is C, and a sequence is as follows: TGAATTAAAATAATCTATGATACCTACGTACGAGTATAAATTTCAACT ATAATAACTAATTTAACGTTCAGACAGTCTCTAATGATTTTTATTCAAAAC TAAATTGTAAGGATATTTTTTTAATCTTTTTTTAATTAAAGAGTAATTTTTT AGGGAACTTCGTAGGGACAAAACCTAGTTGATATTCTAAAATAAAAGAT CAATTATAACCTAAATAAATCACACTTAATTACACCATATTTTTTCAAAAA AGAAAAGAAAAAAGAAAAAAGAAACTTCCTTTTCAATGGGAAACTCCG TCTCTCCTTCCTAACCTCTTTTCCTTTGATTTCATAAACCTTTCTATTCCTT ATATTAAAGATTTCATACACTATTACACGGCCTTTACATTTGTTTCTTTCTC AATATATCTGATTCAGGAAATGCAAATGAATTTTTCTAAGAATCTTCTTTC AAAAAGTAACAAAAATTAGTCTATATATACCCCGCCCTCCCTTTCTACAC AGTTACATATTCAGAACTCTACCAAACTTTCATTTGGCCTTGCTTGAAAG AGTTTCAAACTAACAAAATATGGAGAGACAAAAGAAGAGGGACTTCCA AAATCTTAAGAATGAACATGGTCATGATCGACCTTCTGAAACTAGCCCTA
Description
AAAGGGCATCCAACTCTATTCCAACACCTGTCCGGTCTGCTTCTTGAATC TTCTTCTTTTTTTGTCAAGGATTTTGTGATCGTTAACGATTGTACGAACAC GTGGTGTTCTTGTGTTTCTTTGTTCTCAAGCTGCTCTTCTTCTTCTTCTTC TTTTGCCAAGGATTTTGTGATCGTTAATGATTTTACGAACACGTGGTGTT CTTGTTTTTCTTTGTTTTCAAGCTGCTGATCTTCTTCTTCTTCTTCTTTTGT CAAGAATTTTGTGATCATTAACGATTTTACGAACACGGGGTGCTTTTGTT TTTCTTAGTTCTGAAGCCACGTCCGACATTGTAGCTAACGATTTGTCTCC TCAATTATGGTCTCAAATACAAGATATTCTTCGTGGGCTGGTAATTTTAAT CTCCTTCTCTATCTCTTTTCCAGATCCTATCGTTCTTGGCTATGAATCACTT CTATCCACAACTATTACCGAGCTTCATTTTTGTTATATAACTTTGAGTATCC TACGTAGTGTTTCACTGGGTTCCTCAACCAGACTCTTATCCATCTTTTTAT CTCATTGAAATGGCACTTAGGACAATAAACACCTCT; SEQ ID NO.1. Preferably, the position 600 of the SNP sequence is C. Preferably, the position 600 of the nucleotide sequence has C/T biallelic polymorphism, and a sequence is as follows: TGAATTAAAATAATCTATGATACCTACGTACGAGTATAAATTTCAACT ATAATAACTAATTTAACGTTCAGACAGTCTCTAATGATTTTTATTCAAAAC TAAATTGTAAGGATATTTTTTTAATCTTTTTTTAATTAAAGAGTAATTTTTT AGGGAACTTCGTAGGGACAAAACCTAGTTGATATTCTAAAATAAAAGAT CAATTATAACCTAAATAAATCACACTTAATTACACCATATTTTTTCAAAAA AGAAAAGAAAAAAGAAAAAAGAAACTTCCTTTTCAATGGGAAACTCCG TCTCTCCTTCCTAACCTCTTTTCCTTTGATTTCATAAACCTTTCTATTCCTT ATATTAAAGATTTCATACACTATTACACGGCCTTTACATTTGTTTCTTTCTC AATATATCTGATTCAGGAAATGCAAATGAATTTTTCTAAGAATCTTCTTTC AAAAAGTAACAAAAATTAGTCTATATATACCCCGCCCTCCCTTTCTACAC AGTTACATATTCAGAACTCTACCAAACTTTCATTTGGCCTTGCTTGAAAG AGTTTCAAACTAACAAAATATGGAGAGACAAAAGAAGAGGGACTTYCA AAATCTTAAGAATGAACATGGTCATGATCGACCTTCTGAAACTAGCCCTA AAAGGGCATCCAACTCTATTCCAACACCTGTCCGGTCTGCTTCTTGAATC TTCTTCTTTTTTTGTCAAGGATTTTGTGATCGTTAACGATTGTACGAACAC
Description
GTGGTGTTCTTGTGTTTCTTTGTTCTCAAGCTGCTCTTCTTCTTCTTCTTC TTTTGCCAAGGATTTTGTGATCGTTAATGATTTTACGAACACGTGGTGTT CTTGTTTTTCTTTGTTTTCAAGCTGCTGATCTTCTTCTTCTTCTTCTTTTGT CAAGAATTTTGTGATCATTAACGATTTTACGAACACGGGGTGCTTTTGTT TTTCTTAGTTCTGAAGCCACGTCCGACATTGTAGCTAACGATTTGTCTCC TCAATTATGGTCTCAAATACAAGATATTCTTCGTGGGCTGGTAATTTTAAT CTCCTTCTCTATCTCTTTTCCAGATCCTATCGTTCTTGGCTATGAATCACTT CTATCCACAACTATTACCGAGCTTCATTTTTGTTATATAACTTTGAGTATCC TACGTAGTGTTTCACTGGGTTCCTCAACCAGACTCTTATCCATCTTTTTAT CTCATTGAAATGGCACTTAGGACAATAAACACCTCT; SEQ ID NO.2, wherein Y is T or C, T is thymine deoxyribonucleotide, and C is cytosine deoxyribonucleotide. Preferably, specific identification primers are: SLAF7747-FS: TCGAAATCTTAAGAATGAACATGGT, SEQ ID NO.3; and SLAF7747-FL: CCGAAATCTTAAGAATGAACATGGTCATGAT, SEQ ID NO.4. Preferably, a universal primer is SLAF7747-R: ATCACAAAATCCTTGACA, SEQ ID NO.5. A kit for detecting resistance to cucumber powdery mildews comprises: specific identification primers: SLAF7747-FS: TCGAAATCTTAAGAATGAACATGGT, SEQ ID NO.3; SLAF7747-FL: CCGAAATCTTAAGAATGAACATGGTCATGAT, SEQ ID NO.4; a universal primer: SLAF7747-R: ATCACAAAATCCTTGACA, SEQ ID NO.5. Preferably, specific steps comprise: 1. extracting a genome of a sample to be detected; 2. conducting PCR amplification on the genome; 3. judging disease resistance through an amplification product.
Description
Preferably, a PCR amplification reaction system comprises: reaction buffer 1x; dNTP 0.2mmol/L; Taq DNA synthetase 0.06U/pL; mixed primer 0.8pmol/L; DNA template 100 ng/ul; ddH20 balance. Preferably, by the mass of matters, SLAF7747-FS: SLAF7747-FL: SLAF7747-R=1:1:1. A fragment length difference of the PCR amplification product is judged and detected through polypropylene gel electrophoresis or capillary electrophoresis. To sum up, the present invention discloses the SNP molecular marker site for the resistance to the cucumber powdery mildews and the application thereof. By selecting a breeding material by virtue of the molecular marker technology at DNA level, rapid identification of a cucumber germplasm material can be realized, so that an offspring material with target genes is rapidly selected to solve the problems of long identification time for disease resistance and difficulty in restoration after identification. The present invention is simple, feasible and high in efficiency and is an effective measure of identifying a cucumber material resisting the powdery mildews for conventional crossing and backcrossing methods.
Description of Drawings
Fig. 1 is an actual detection result of a field sample. Lane 1 is Marker; lane 2 is a disease-resistant plant, lane 3 is a susceptible plant, and lanes 4-15 are plants to be detected.
Detailed Description
The technical solutions in the embodiments of the present invention are described clearly and completely below. Apparently, the described embodiments
Description
are only part rather than all of the embodiments of the present invention. On the basis of the embodiments in the present invention, all other embodiments obtained by those ordinary skilled in the art without contributing creative effort belong to the protection scope of the present invention. Embodiment 1 Construction of plant material: With disease resistant materials BK2 as male parents to serve as disease resistant controls, and susceptible plants H136 as female parents to serve as susceptible controls and samples to be detected, disease resistant indexes were analyzed according to disease resistant levels of the materials after powdery mildews were inoculated to the materials. All the materials were planted in a Yangdu test site in Zhejiang Academy of Agricultural Sciences. Inoculation and phenotype identification of powdery mildews Parents BK2, H136 and the samples to be detected were potted in an intelligent greenhouse. In the one-leaf-and-one-leaflet stage of seedlings of cucumbers, a spray inoculation method was employed, wherein an inoculation concentration was 1x1O spores/niL, a day temperature of an environment temperature was 28°C, a night temperature was 18°C, and environment humidity was 85%. The incidence was observed and recorded at day 7 after inoculation. Identification of resistant molecules Disease resistant materials (disease resistant controls), susceptible materials (susceptible controls) and 12 samples to be detected were selected and planted, 0.1g of tender leaves of cucumbers in the two-leaf-and-one-leaflet stage were taken, and genomic DNA of the cucumbers was extracted by using an SDS method and diluted to 60ng/pL. PCR amplification was conducted by using the following primers: forward primers: SLAF7747-FS: TCGAAATCTTAAGAATGAACATGGT, SEQ ID NO.3; SLAF7747-FL: CCGAAATCTTAAGAATGAACATGGTCATGAT, SEQ ID NO.4;
Description
a reverse primer: SLAF7747-R: ATCACAAAATCCTTGACA, SEQ ID NO.5. 50pL system: reaction buffer 1x; dNTP 0.2mmol/L; Taq DNA synthetase 0.06U/pL; mixed primer 0.8pmol/L (by the mass of matters, SLAF7747-FS: SLAF7747-FL: SLAF7747-R=1:1:1); DNA template 100 ng/ul; ddH20 balance. The reaction system was pre-denatured for 4 min at 94°C, denatured for 1 min at 94°C, annealed for s at 56°C, extended for 30s at 72°C, cycled for 30 times, extended for 5 min at 72°C and preserved at 4°C. The PCR amplification products were subjected to capillary electrophoresis. Results were shown in Fig. 1. PCR amplification products of the disease resistant materials (disease resistant controls) move faster than amplification products of the susceptible materials (susceptible controls). After capillary electrophoresis, disease resistant amplification bands were lower than susceptible amplification bands. In 12 samples to be detected, sample 1, sample 2, sample 4, sample 5, sample 7, sample 9 and sample 10 were susceptible plants; sample 3, sample 6 and sample 12 were disease resistant plants; and sample 8 and sample 11 were moderate resistance. Resistance phenotype identification: Positive controls, negative controls and the samples to be detected were potted in the intelligent greenhouse. In the one-leaf-and-one-leaflet stage of seedlings of cucumbers, the spray inoculation method was employed, wherein an inoculation concentration was 1x1O' spores/mL. The incidence was observed and recorded at day 7 after inoculation. Powdery mildew resistant/susceptible phenotypic results were consistent to results of electrophoresis (Fig. 1). It can be
Description
seen that molecular identification results were consistent with the actual results and can be used for agricultural production. Each embodiment in the description is described in a progressive way. The difference of each embodiment from each other is the focus of explanation. The same and similar parts among all of the embodiments can be referred to each other. The above description of the disclosed embodiments enables those skilled in the art to realize or use the present invention. Many modifications to the above embodiments will be apparent to those skilled in the art. The general principle defined herein can be realized in other embodiments without departing from the spirit or scope of the present invention. Therefore, the present invention will not be limited to these embodiments shown herein, but will conform to the widest scope consistent with the principle and novel features disclosed herein.
Claims (7)
1. An application of an SNP molecular marker for resistance to cucumber powdery mildews in preparing a formulation for detecting resistance to the cucumber powdery mildews, wherein position 600 of a nucleotide sequence of the SNP molecular marker is C, and a sequence is as follows: TGAATTAAAATAATCTATGATACCTACGTACGAGTATAAATTTCAACT ATAATAACTAATTTAACGTTCAGACAGTCTCTAATGATTTTTATTCAAAAC TAAATTGTAAGGATATTTTTTTAATCTTTTTTTAATTAAAGAGTAATTTTTT AGGGAACTTCGTAGGGACAAAACCTAGTTGATATTCTAAAATAAAAGAT CAATTATAACCTAAATAAATCACACTTAATTACACCATATTTTTTCAAAAA AGAAAAGAAAAAAGAAAAAAGAAACTTCCTTTTCAATGGGAAACTCCG TCTCTCCTTCCTAACCTCTTTTCCTTTGATTTCATAAACCTTTCTATTCCTT ATATTAAAGATTTCATACACTATTACACGGCCTTTACATTTGTTTCTTTCTC AATATATCTGATTCAGGAAATGCAAATGAATTTTTCTAAGAATCTTCTTTC AAAAAGTAACAAAAATTAGTCTATATATACCCCGCCCTCCCTTTCTACAC AGTTACATATTCAGAACTCTACCAAACTTTCATTTGGCCTTGCTTGAAAG AGTTTCAAACTAACAAAATATGGAGAGACAAAAGAAGAGGGACTTCCA AAATCTTAAGAATGAACATGGTCATGATCGACCTTCTGAAACTAGCCCTA AAAGGGCATCCAACTCTATTCCAACACCTGTCCGGTCTGCTTCTTGAATC TTCTTCTTTTTTTGTCAAGGATTTTGTGATCGTTAACGATTGTACGAACAC GTGGTGTTCTTGTGTTTCTTTGTTCTCAAGCTGCTCTTCTTCTTCTTCTTC TTTTGCCAAGGATTTTGTGATCGTTAATGATTTTACGAACACGTGGTGTT CTTGTTTTTCTTTGTTTTCAAGCTGCTGATCTTCTTCTTCTTCTTCTTTTGT CAAGAATTTTGTGATCATTAACGATTTTACGAACACGGGGTGCTTTTGTT TTTCTTAGTTCTGAAGCCACGTCCGACATTGTAGCTAACGATTTGTCTCC TCAATTATGGTCTCAAATACAAGATATTCTTCGTGGGCTGGTAATTTTAAT CTCCTTCTCTATCTCTTTTCCAGATCCTATCGTTCTTGGCTATGAATCACTT CTATCCACAACTATTACCGAGCTTCATTTTTGTTATATAACTTTGAGTATCC TACGTAGTGTTTCACTGGGTTCCTCAACCAGACTCTTATCCATCTTTTTAT CTCATTGAAATGGCACTTAGGACAATAAACACCTCT; SEQ ID NO.1.
Claims
2. The application of the SNP molecular marker for resistance to cucumber powdery mildews in preparing the formulation for detecting resistance to the cucumber powdery mildews according to claim 1, wherein the position 600 of the nucleotide sequence has C/T biallelic polymorphism, and a sequence is as follows: TGAATTAAAATAATCTATGATACCTACGTACGAGTATAAATTTCAACT ATAATAACTAATTTAACGTTCAGACAGTCTCTAATGATTTTTATTCAAAAC TAAATTGTAAGGATATTTTTTTAATCTTTTTTTAATTAAAGAGTAATTTTTT AGGGAACTTCGTAGGGACAAAACCTAGTTGATATTCTAAAATAAAAGAT CAATTATAACCTAAATAAATCACACTTAATTACACCATATTTTTTCAAAAA AGAAAAGAAAAAAGAAAAAAGAAACTTCCTTTTCAATGGGAAACTCCG TCTCTCCTTCCTAACCTCTTTTCCTTTGATTTCATAAACCTTTCTATTCCTT ATATTAAAGATTTCATACACTATTACACGGCCTTTACATTTGTTTCTTTCTC AATATATCTGATTCAGGAAATGCAAATGAATTTTTCTAAGAATCTTCTTTC AAAAAGTAACAAAAATTAGTCTATATATACCCCGCCCTCCCTTTCTACAC AGTTACATATTCAGAACTCTACCAAACTTTCATTTGGCCTTGCTTGAAAG AGTTTCAAACTAACAAAATATGGAGAGACAAAAGAAGAGGGACTTYCA AAATCTTAAGAATGAACATGGTCATGATCGACCTTCTGAAACTAGCCCTA AAAGGGCATCCAACTCTATTCCAACACCTGTCCGGTCTGCTTCTTGAATC TTCTTCTTTTTTTGTCAAGGATTTTGTGATCGTTAACGATTGTACGAACAC GTGGTGTTCTTGTGTTTCTTTGTTCTCAAGCTGCTCTTCTTCTTCTTCTTC TTTTGCCAAGGATTTTGTGATCGTTAATGATTTTACGAACACGTGGTGTT CTTGTTTTTCTTTGTTTTCAAGCTGCTGATCTTCTTCTTCTTCTTCTTTTGT CAAGAATTTTGTGATCATTAACGATTTTACGAACACGGGGTGCTTTTGTT TTTCTTAGTTCTGAAGCCACGTCCGACATTGTAGCTAACGATTTGTCTCC TCAATTATGGTCTCAAATACAAGATATTCTTCGTGGGCTGGTAATTTTAAT CTCCTTCTCTATCTCTTTTCCAGATCCTATCGTTCTTGGCTATGAATCACTT CTATCCACAACTATTACCGAGCTTCATTTTTGTTATATAACTTTGAGTATCC TACGTAGTGTTTCACTGGGTTCCTCAACCAGACTCTTATCCATCTTTTTAT CTCATTGAAATGGCACTTAGGACAATAAACACCTCT; SEQ ID NO.2.
Claims
3. The application of the SNP molecular marker for resistance to cucumber powdery mildews in preparing the formulation for detecting resistance to the cucumber powdery mildews according to claim 2, wherein specific identification primers are: SLAF7747-FS: TCGAAATCTTAAGAATGAACATGGT, SEQ ID NO.3; and SLAF7747-FL: CCGAAATCTTAAGAATGAACATGGTCATGAT, SEQ ID NO.4.
4. A kit for detecting resistance to cucumber powdery mildews, comprising: specific identification primers: SLAF7747-FS: TCGAAATCTTAAGAATGAACATGGT, SEQ ID NO.3; SLAF7747-FL: CCGAAATCTTAAGAATGAACATGGTCATGAT, SEQ ID NO.4; a universal primer: SLAF7747-R: ATCACAAAATCCTTGACA, SEQ ID NO.5.
5. A use method of the kit for detecting resistance to cucumber powdery mildews of claim 4, comprising specific steps: 1. extracting a genome of a sample to be detected; 2. conducting PCR amplification on the genome; 3. judging disease resistance through differences of an amplification product.
6. The use method according to claim 5, wherein a PCR amplification reaction system comprises: reaction buffer 1x; dNTP 0.2mmol/L; Taq DNA synthetase 0.06U/pL; mixed primer 0.8pmol/L; DNA template 100 ng/ul; ddH20 balance.
7. The use method according to claim 6, wherein in the mixed primer, by the mass of matters, SLAF7747-FS: SLAF7747-FL: SLAF7747-R=:1:1.
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| AU (1) | AU2021102786A4 (en) |
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2021
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