Method for acquiring chicken whole genome high-density SNP marker sites

A technology of whole genome and marker loci, which is applied in the field of obtaining high-density SNP marker loci in the whole genome of chickens, can solve the problems of interfering experimental results, affecting typing efficiency and cost, data loss, etc., and achieves technical stability and high repeatability Effect

Active Publication Date: 2016-01-13
CHINA AGRI UNIV
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Problems solved by technology

However, different simplified genome sequencing methods are quite different in terms of library construction strategy, single-enzyme digestion/double-enzyme digestion combination selection, and sequencing platform selection, which will significantly affect the efficiency and cost of subsequent typing
For example, the library construction strategy of the RAD sequencing method is complicated, and too many steps will interfere with the results of subsequent experiments; the frequency and distribution of different restriction endonucleases on the genomes of different species are quite different. Species, which enzyme to use for experiments will ...

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  • Method for acquiring chicken whole genome high-density SNP marker sites
  • Method for acquiring chicken whole genome high-density SNP marker sites
  • Method for acquiring chicken whole genome high-density SNP marker sites

Examples

Experimental program
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Embodiment 1

[0041] Embodiment 1 is used to illustrate the method of the present invention

[0042] 1. Experimental materials:

[0043] Collection of red native chickens, commercial layer breeds Bailaihang chickens, commercial broiler breeds Aiba Yijia broiler chickens, Lingnan yellow chickens, Chinese local chicken breeds Huiyang bearded chickens, Wenchang chickens, Henan game cocks, Qingyuan Ma chickens, black wolf pheasant chickens, Camellia Blood samples from 4-6 individual chickens, Peking chicken, Tibetan chicken, silky chicken, Shouguang chicken, bamboo silk chicken, Shiqi mixed chicken, Xianju chicken, invisible white chicken, and bantam yellow chicken, totaling 96 Individuals, the genome was extracted, and the genome concentration was diluted to 50ng / μL for use.

[0044] 2. Adapter and primer sequence:

[0045] Synthesize a pair of universal adapter sequences, 96 pairs of barcode adapter sequences, and a pair of PCR primer sequences.

[0046] 3. Sequencing library construction:...

Embodiment 2

[0068] The selection of embodiment 2 chicken genome optimum endonuclease combinations

[0069] Example 2 is used to illustrate the enzyme cleavage combination used in the present invention.

[0070] Considering the recognition characteristics of different restriction enzyme sites (such as the number of recognized bases, GC content, methylation status), etc., the inventor designed a total of 8 sets of double enzyme digestion combinations, and carried out different enzyme digestion through 3 individuals of Lingnan yellow chicken and Huiyang bearded chicken. The combined sequencing experiment, the experimental process is the same as that in Example 1, and the experimental results are shown in Table 2. It can be seen that the number of SNPs in the EcoRI–MseI digestion combination is 134,291 (the number of SNPs will vary with the number of experimental individuals), and the number of enzyme-digested fragments is 414,294, with the highest comparison rate with the genome. In one Next...

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Abstract

The invention belongs to the technical field of genetic engineering, and provides a method for acquiring chicken whole genome high-density SNP marker sites. The method comprises the following steps: (1) predicting distribution of restriction fragments acquired from double-restriction chicken genomes of EcoRI and MseI; (2) designing a universal joint, a barcode joint and a PCR amplification primer according to the distribution characteristics of the restriction fragments of EcoRI and MseI; (3) constructing a simplified genome sequencing library; (4) sequencing by virtue of the library constructed in the step (3) through a computer; and (5) according to a sequencing result, acquiring SNP marker sites. The method disclosed by the invention provides a universal strategy for the construction of a whole genome high-density SNP map by virtue of double-restriction GBS for different varieties of chickens, so that the cost on acquiring every SNP marker site is reduced by an order of magnitude compared with a conventional chip technology; and the method is stable in technology and high in repeatability.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for obtaining high-density SNP marker sites in the whole genome of chickens based on sequencing genotyping technology. Background technique [0002] As a model organism of poultry, the chicken became the first agricultural economic animal to complete complete gene sequencing in 2004. Due to the huge biodiversity of different chicken breeds, it is being used more and more as a high-quality genetic model To quantitative genetics and molecular breeding, functional gene mapping, gene regulation and development and other fields. Molecular markers are an important tool for studying biological genetic variation. Single nucleotide polymorphisms (SNP), as the third generation of molecular markers, have the characteristics of large number, wide distribution, and genetic stability, and are widely used in linkage analysis, global Genetic breeding fields such as genome association analy...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCC12Q1/6827C12Q1/6869
Inventor 胡晓湘王宇哲曹学敏李宁
Owner CHINA AGRI UNIV
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