Genetic marker using pig CKM (creatine kinase muscle) 5' flanking promoter region SNP (single nucleotide polymorphism) as pig carcass traits and application

A technology of genetic markers and pig carcasses, applied in the direction of DNA / RNA fragments, recombinant DNA technology, etc., can solve the problems of unrevealed relationship between pig carcass traits

Inactive Publication Date: 2014-04-09
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, so far, there has been no relevant report on the SNP of the pig CKM gene, and its relationship with pig carcass and other traits has not been revealed.

Method used

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  • Genetic marker using pig CKM (creatine kinase muscle) 5' flanking promoter region SNP (single nucleotide polymorphism) as pig carcass traits and application
  • Genetic marker using pig CKM (creatine kinase muscle) 5' flanking promoter region SNP (single nucleotide polymorphism) as pig carcass traits and application
  • Genetic marker using pig CKM (creatine kinase muscle) 5' flanking promoter region SNP (single nucleotide polymorphism) as pig carcass traits and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Example 1 Obtaining specific DNA fragments of the 5' flanking promoter region of pig CKM and establishing a SNP detection method

[0023] Two pig breeds with different genotypes, the foreign blood-related pig "Large White pig" and the Chinese local pig breed "Meishan pig", were selected as test materials, and the following primers were designed according to the genome sequence of the pig CKM gene (GeneBank accession number DQ153192). The primer sequences are as follows:

[0024] Forward primer F: 5'AGGAGACAGCGAGTAGCG 3',

[0025] Reverse primer R: 5'CCCTGATGCCTGTGCTTA3'.

[0026] Use the above primers to carry out PCR amplification in large white pig (foreign blood-related pig) and Meishan pig (Chinese local pig) genomic DNA (for the extraction method of pig genomic DNA, please refer to the literature: Xiong Yuanzhu. Introduction to the Biochemical and Molecular Genetic Experiments of Pigs. Beijing , China Agricultural Press, 1999, 39-51).

[0027] The PCR reaction sy...

Embodiment 2

[0035] Example 2: Association analysis and application of genetic markers cloned in the present invention and pig carcass traits

[0036] In order to determine whether the SNP in the 5' flanking promoter region of pig CKM gene is related to the difference of pig phenotype, 135 Large White pigs × Meishan pig F2 generation established by the Key Open Laboratory of Pig Genetics and Breeding, Ministry of Agriculture, Huazhong Agricultural University, Wuhan City, Hubei Province, China were selected. The resource groups are experimental materials (XuDQ, Xiong YZ, Liu M, Lan J, Ling XF, Deng CY, Jiang SW. Association analyzes with carcass traits in the porcine KIAA 1717 and HUMMLC2B genes. Asian-Aust J Anim Sci. 2005, 18: 1519-1523), using PCR-DraIII-RFLP method (Deng GR.A sensitive non-radioactive PCR-RFLP analysis for detecting point mutations at 12th codon of oncogene c-Ha-ras in DNAs of gastric cancer. Nucleic Acids Res.1988, 16:6231) for polymorphism detection, and analyzed the ...

Embodiment 3

[0043] Embodiment 3 The expression level analysis of the CKM gene of different genotypes of the present invention

[0044] Extract the DNA and RNA (see Invitrogen) from the 2-month-old muscles of Meishan pigs and Large White pigs (Xiong Yuan. Introduction to the Biochemical and Molecular Genetic Experiments of Pigs. Beijing, China Agricultural Press, 1999, 39-51) Reagent instructions), and reverse transcribe the RNA into cDNA. The specific DNA fragment of the 5' flanking promoter region of the CKM gene was amplified by PCR using pig genomic DNA as a template, and the amplified fragment was digested with DraIII to obtain different genotypes. GG type. Using the reverse-transcribed cDNA as a template, the designed primers were used for real-time quantitative PCR to analyze the expression levels of different genotypes.

[0045] The DNA sequences of the primers used in real-time quantitative PCR are as follows:

[0046] Forward primer F: TCACCTGCCCGTCTAACCT,

[0047] Reverse p...

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Abstract

The invention belongs to the technical field of pig molecular marker preparation and particularly relates to a genetic marker using pig CKM (creatine kinase muscle) 5' flanking promoter segments as pig carcass traits and application. The genetic marker has the nucleotide sequence shown as SEQ ID NO (sequencer identifier number):1 and SEQ ID NO:2 in a sequence table. The genetic marker is characterized in that A / G and G / A mutations respectively exist at the 82bp part of the sequence, and the polymorphism of DraIII-RFLP is caused. The genetic marker is used for carrying out pig carcass traits correlation analysis on large white pig and meishan pig F2 generations, and the expression quantity of CKM in muscular tissues of large white pigs and meishan pigs containing different genotypes is detected. The invention also discloses a parting detection method of the genetic marker, and the novel genetic marker and the detection method are provided for the pig carcass traits molecular marker auxiliary selection.

Description

technical field [0001] The invention relates to the fields of pig breeding and molecular marker-assisted selection of pigs, in particular to the cloning of a CKM 5' flank promoter fragment of a gene related to pig carcass traits and its application as a genetic marker for pig carcass traits. Background technique [0002] In recent years, with the rapid development of genome research and the continuous deepening of gene structure and function research, a new type of selection method that can use DNA polymorphisms as markers for genetic analysis at the molecular level to identify individual genotype differences—molecular markers Assisted selection technology came into being. Molecular markers are directly expressed in the form of DNA, which can be detected in all tissues and developmental stages of organisms, and are not restricted by seasonal environments; polymorphisms are abundant and large in number; most of them are co-dominant, and can distinguish homozygous and heterozy...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/113C12N15/11C12Q1/68
Inventor 徐德全孙小瑞刘敏熊远著邓昌彦蒋思文
Owner HUAZHONG AGRI UNIV
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