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Cotton whole genome 40K mononucleotide site, and application thereof in cotton genotyping

A genome-wide, single-nucleotide technology, applied in the fields of genetics, bioinformatics, cotton molecular breeding, and molecular biology, can solve problems such as no chip reports

Active Publication Date: 2021-08-27
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] Single-nucleotide liquid-phase gene chips based on GBTS technology have been developed and screened in corn, rice, and soybean, but there are no reports of such chips in cotton

Method used

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  • Cotton whole genome 40K mononucleotide site, and application thereof in cotton genotyping
  • Cotton whole genome 40K mononucleotide site, and application thereof in cotton genotyping
  • Cotton whole genome 40K mononucleotide site, and application thereof in cotton genotyping

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Example 1. Cotton 40K single nucleotide loci are used in the preparation of cotton gene chips.

[0029] The specific method for the preparation of the 40K cotton gene chip is: through library construction, hybrid capture, library quality inspection and sequencing analysis, the 40K target loci are finally determined.

[0030] 1. Library Construction of 13 Cotton Varieties

[0031] Add 100 ng of cotton DNA, 4 μL of GenoBaits End Repair Buffer, 3.1 μL of GenoBaits End repair Enzyme to a 0.2 μL PCR tube, and make up 20 μL with water; place the reaction tube in a PCR machine, at 37°C for 20 minutes, and at 72°C for 20 minutes. Then add 2 μL of GenoBaits Ultra DNA ligase, 8 μL of GenoBaits Ultra DNA LigaseBuffer, 2 μL of GenoBaits Adapter, 8 μL of Nuclease-free water, and place in a PCR machine at 22°C for 60 min. Add 1.2 times of GenoPrep DNA Clean Beads, mix thoroughly, place on a magnetic stand, let stand for 3min, and remove the supernatant. Add 100 μL of freshly prepar...

Embodiment 2

[0180] Example 2. Application of cotton 40K single nucleotide liquid-phase gene chip in cotton genotyping.

[0181] The inventor selected a total of 13 tetraploid cotton species materials including the genetic standard line TM-1 of upland cotton, American cotton, semi-wild cotton, the backbone parents of the three cotton regions of the Yangtze River Basin, the Yellow River Basin, and the Northwest Inland of my country. The 40K cotton gene chip prepared in Example 1 detects the genotypes of the 13 variety materials at the 40K single nucleotide locus described in Example 1. The detection rate of 13 kinds of material loci was 98.62~99.96%. By comparing the loci of the reference genome (TM-1) with the loci detected by the chip, the consistency of the detected loci was 99.40%.

[0182] Attached Table 2, 13 Cotton Variety Resources

[0183]

[0184]

[0185] The entire experimental process is completed based on the GenoBaits technology system. The specific experimental proce...

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Abstract

The invention provides a cotton whole genome 40K mononucleotide site, and application thereof in cotton genotyping. The cotton whole genome 40K mononucleotide site is obtained through a VCF file obtained based on three-population re-sequencing, a cotton genome TM-1V_2.1 is taken as a reference, a target segment is preliminarily screened from site information in data, and then the probe corresponding to the target segment is synthesized; 13 cotton varieties are selected for data testing, and 13 upland cotton materials including one wild species, one semi-wild species and 11 cultivated species are selected for chip detection; according to a detection result, a target area with good uniformity is preferentially selected, and overall uniform distribution is ensured; and finally screening is performed to obtain the 40K mononucleotide site. The application of the site can greatly improve the genotype detection accuracy, improve the detection level and reduce the detection cost, and provides important technical support for molecular breeding of cotton.

Description

technical field [0001] The invention relates to the fields of genetics, molecular biology, bioinformatics and cotton molecular breeding, and more specifically relates to the 40K single nucleotide site of the whole cotton gene and its application in cotton genotyping. Background technique [0002] Molecular detection is an important means of genetic variation analysis. Ideal molecular testing would be complete testing of organisms at the individual and population levels, such as whole-genome sequencing. Although the cost of detection has decreased exponentially with the development of molecular detection technology, the full-information molecular detection of organisms still faces major challenges in information processing and analysis. Therefore, the genetic variation detection technology based on molecular markers is still a simple, efficient and low-cost molecular detection method. Molecular marker technology is to use each marker distributed on the biological genome as ...

Claims

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Application Information

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IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/13C12Q2600/156
Inventor 胡艳司占峰韩泽刚方磊张天真
Owner ZHEJIANG UNIV
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