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Core SNP markers for identification of cotton hybrids developed based on KASP technology

A hybrid and marker technology, applied in the field of molecular biology, can solve the problems of few detection sites, poor representativeness of results, unsuitable for data sharing, etc.

Active Publication Date: 2019-01-04
INST OF COTTON RES CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in the course of practice, it has also exposed its insurmountable defects such as fewer detection sites, poor representativeness of results, and unsuitable data sharing.

Method used

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  • Core SNP markers for identification of cotton hybrids developed based on KASP technology
  • Core SNP markers for identification of cotton hybrids developed based on KASP technology
  • Core SNP markers for identification of cotton hybrids developed based on KASP technology

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Example 1 26 core SNP markers for cotton hybrid identification developed based on KASP technology

[0028] Based on the whole genome sequence of the upland cotton genetic standard line TM-1, the version number is NBI G.hirsutum genome (https: / / www.cottongen.org / ) to screen and locate SNP loci.

[0029] The screening and determination of the core SNP sites in the present invention: based on sample data with a wide range of representative genetic backgrounds, using 63k Illumina cotton genome-wide SNP chips, screening and evaluating according to the following characteristics: (a) whether it is a unit site; (b) Whether the locus evaluation score GenTrain score is above 0.6; (c) whether the locus polymorphism and the minimum allele frequency are above 0.2; (d) the distribution on the genome; (e) the homozygous rate of hybrid parents Hybrid F 1 Heterozygosity rate; (f) Whether the clusters of the three genotypes are easy to distinguish and other factors. Each site was screened an...

Embodiment 2

[0037] Example 2 Using core SNP markers to detect the purity of the cotton hybrid'Zhongmianzuo 63'

[0038] The detection method is as follows:

[0039] 1. DNA extraction

[0040] Randomly pick 96 seeds of the hybrid seed of the "Zhongmian Institute 63" to be tested for DNA preparation.

[0041] (1) Dehull the single cotton seed, pulverize it and transfer it to a 2ml centrifuge tube.

[0042] (2) Add 800μl DNA extraction solution (1% SDS, 0.01mol / L EDTA 8.0, 0.7mol / L NaCl, 0.05mol / L Tris-HCl, 0.5% sorbitol, 1% PVP, 1% β-mercaptoethanol) , After vortexing to fully mix, water bath at 65℃ for 30min, shaking gently at intervals of about 10min.

[0043] (3) After the end of the water bath, add an equal volume of 800 μl of a mixture of phenol, chloroform and isoamyl alcohol (volume ratio 25:24:1), mix upside down until there is no layering, and centrifuge at 10,000 rpm for 10 min.

[0044] (4) Transfer the supernatant to another 2ml centrifuge tube, add 0.7 times the volume of isopropanol and ...

Embodiment 3

[0055] Example 3 Using core SNP markers to identify whether the cotton species to be tested is a hybrid species "Zhongmianso 72"

[0056] The detection method is as follows:

[0057] Randomly select the cotton seeds to be tested and 24 seeds of the'China Cotton Institute 72' standard sample seeds, and perform DNA preparation according to the steps in Example 2, and use 26 core SNP markers for genotyping. The PCR reaction was the same as that described in Example 2.

[0058] Result analysis: Through the above detection, the typing results of the tested cotton species and the'China Cotton Institute 72' at 26 SNP loci were obtained. The comparison analysis results showed that the tested hybrids were in the 16 SNP loci and Cotton Institute 72' showed a difference. It shows that the tested hybrid is not a true hybrid of'China Cotton Institute 72'.

[0059] As there are many detection methods for SNP markers, the specific conditions are divided according to the detection throughput as fol...

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Abstract

The invention provides core SNP markers developed based on KASP (competitive allele specific) technology and applied to cotton hybrid identification. The SNP markers are distributed on 26 chromosomes of tetraploid genome of cotton, and every SNP marker is located on each chromosome, totaling 26 SNP markers. On the basis of the set of core SNP markers, high-throughput SNP typing detection on cotton hybrid is achieved.

Description

Technical field [0001] The invention relates to the field of molecular biology, in particular to a core SNP marker for cotton hybrid identification developed based on KASP technology. Background technique [0002] Cotton is the main economic crop in my country, and the cultivation and promotion of good new varieties is of great significance to the development of the national economy and the improvement of people's living standards. In recent years, with the rapid development of cotton hybrid breeding and the heterosis shown by hybrids, cotton hybrids have been widely promoted and planted in my country. Cotton hybrids have excellent characteristics such as high yield, high quality, and strong stress resistance, which generally increase the yield by about 15% compared with conventional varieties. However, the production of cotton hybrid seeds is a labor-intensive industry, and the cost of seed production is relatively high. The purchase price of hybrid seed cotton is generally 4-5...

Claims

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Application Information

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IPC IPC(8): C12N15/11C12Q1/6895
Inventor 匡猛魏守军杨伟华王延琴周大云马磊方丹徐双娇单峰
Owner INST OF COTTON RES CHINESE ACAD OF AGRI SCI
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