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Method for screening molecular markers correlative with properties based on sequencing technique and BSA (Bulked Segregant Analysis) technique

A sequencing and labeling technology, applied in biochemical equipment and methods, microbial measurement/inspection, etc., can solve the problems of high price, low accuracy of marker and trait association analysis, low throughput, etc., and achieve development cost reduction, Effects of Marker Development Throughput Improvement and Complexity Reduction

Active Publication Date: 2013-09-18
BIOMARKER TECH
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Problems solved by technology

[0004] The purpose of the present invention is to provide a trait-related molecular marker screening method based on sequencing and BSA technology. Applied to marker development and gene positioning in crop molecular breeding, etc.

Method used

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  • Method for screening molecular markers correlative with properties based on sequencing technique and BSA (Bulked Segregant Analysis) technique
  • Method for screening molecular markers correlative with properties based on sequencing technique and BSA (Bulked Segregant Analysis) technique
  • Method for screening molecular markers correlative with properties based on sequencing technique and BSA (Bulked Segregant Analysis) technique

Examples

Experimental program
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Effect test

Embodiment 1

[0027] Example 1 Population-specific Length Amplified Fragment Sequencing Detection of DNA Molecular Markers

[0028] see figure 1 shown, including the following steps:

[0029] (1) The objects processed in this embodiment are the DNA samples of the two parents and the DNA samples of the isolated populations after identification; the individual samples in each isolated population should be mixed in the same proportion.

[0030] (2) In this embodiment, the genome or BAC reference sequence of the species to be studied is analyzed, the training parameters are used to find the optimal enzyme combination, and the optimal length of the amplified product is selected to meet the following conditions:

[0031] aThe restriction sites of the enzyme are evenly distributed on the genome;

[0032] b Selecting a specific length of restriction fragments can ensure the number of sequence tags;

[0033] c Select a specific length of restriction fragments to avoid falling in highly repetitive...

Embodiment 2

[0043] Example 2 Development of high-density markers and rapid mapping of male sterility trait genes in Chinese cabbage based on high-throughput sequencing

[0044] Including the following steps:

[0045] 1. If Figure 3A Shown is the density distribution map of the 450-500bp length digestion fragments on the chromosome after simulating Ecor1+Msel two enzymes to digest the Chinese cabbage genome reference sequence. After analysis, the 450-500bp length fragment with the highest efficiency is selected, and the length fragment can be evenly distributed on all chromosomes of the genome.

[0046] 2. If Figure 3B Shown is the distribution of the number of 450-500bp length fragments per 100K of the whole genome after simulating Ecor1+Msel two enzymes to digest the Chinese cabbage genome reference sequence. The density of the length segment can also meet the design requirements.

[0047] 3. Through the above analysis, the enzyme digestion parameters of cabbage genome are evaluate...

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Abstract

The invention provides a method for screening DNA molecular markers closely correlative with properties, which is mainly used for identifying DNA molecular markers correlative with properties by combining super bulked segregant analysis (BSA) with a high throughput sequencing technique. The method comprises the following steps of: carrying out parameter training to data comprising different species genomes or BAC (Bacterial Artificial Chromosome) sequences and the like by utilizing a bioinformatics method to find out an optimal marker sequencing scheme for improving the development efficiency of the markers; carrying out correlative analysis to bulked segregant sequencing results to detect the molecular markers correlative with the properties and position back to the genomes or BAC, and then precisely positioning candidate functional genes through the high-density property-correlative molecular markers. Compared with the traditional method, the method has the advantages of greatly improving throughput and greatly reducing cost. The method can be used for carrying out marker positioning to large groups and directly determining the linkage between the markers and the target properties, has high reliability and accuracy of correlative analysis, and is mainly applied to the aspects of marker development and gene positioning, such as crop molecular breeding and the like.

Description

technical field [0001] The present invention relates to a trait-related molecular marker screening method based on sequencing and BSA technology, in particular, it relates to a combination of super BSA and high-throughput sequencing technology to perform deep sequencing on large populations by reducing the complexity of the genome, and A method for identifying trait-associated DNA molecular markers by association analysis. Background technique [0002] The current molecular markers are mainly divided into three categories: the first category is based on electrophoresis and molecular hybridization technology such as RFLP technology; the second category is based on electrophoresis and PCR technology such as AFLP technology; the third category is based on DNA chips and molecular markers of sequencing technologies such as SNPs. Among them, the throughput of polymorphisms detected by the first two methods is small, and the third method is more expensive. [0003] With the devel...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68
Inventor 王晓武郑洪坤
Owner BIOMARKER TECH
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