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Method for detecting single nucleotide polymorphism of goat signal transducer and activator of transcription (STAT3) gene and application of STAT3 gene

A single nucleotide polymorphism, gene technology, applied in biochemical equipment and methods, microbial determination/inspection, etc., can solve the problem of lack of functional research of gene loci, blank and other problems

Inactive Publication Date: 2014-06-18
NORTHWEST A & F UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There is a lack of research on the genetic variation of the Chinese local goat STAT3 gene, and the functional research on this gene locus is even more blank

Method used

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  • Method for detecting single nucleotide polymorphism of goat signal transducer and activator of transcription (STAT3) gene and application of STAT3 gene
  • Method for detecting single nucleotide polymorphism of goat signal transducer and activator of transcription (STAT3) gene and application of STAT3 gene
  • Method for detecting single nucleotide polymorphism of goat signal transducer and activator of transcription (STAT3) gene and application of STAT3 gene

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0066] Embodiment 1, goat STAT3 gene contains the design of 45204, 62058 and 62230 PCR primers

[0067] Since cattle and sheep are both ruminants and highly homologous, and the sequence of the goat genome has not been released so far, the sequence of the bovine STAT3 gene can only be searched on NCBI, and Primer5. 2, 10 and 11 intronic PCR primers for SNP sites, the primer sequences are as follows:

[0068] P1: Upstream primer: 5'-TGTGTGAGTGTGTAGGTTTCAGAAT-3'(25bp),

[0069] Downstream primer: 5'-AGATAGGTTATTAATTAATCAACT A-3'(26bp);

[0070] P2: Upstream primer: 5'-AAATCCAGCCTGAGGGAGAATTCCT-3'(25bp),

[0071] Downstream primer: 5'-GAGTCTCTTTGAAAGTCCACTTTG A-3'(26bp);

[0072] P3: Upstream primer: 5'-TGGGAAAGATACTTGCTG-3'(18bp),

[0073] Downstream primer: 5'-GACCTGAATCACAGGAGGAAAAGA C-3'(26bp)

[0074] Using the above primer pair P1 to amplify the goat genome, it is possible to amplify a 290bp fragment including the 44941nt-45230nt in the second intron region of the...

Embodiment 2

[0083] Embodiment 2, carry out PCR amplification STAT3 gene fragment of goat to be tested with primer P

[0084] 1. Collection of goat samples

[0085] The animals used in the experiment were a total of 585 samples of two breeds, of which

[0086] 1) 220 samples of Hainan Black Goat (HNBG) were collected from the Hainan Black Goat Breeding Farm in Zhanzhou, Hainan Province. Random sampling method was used to collect individual ear tissue samples from the Hainan Black Goat Breeding Farm in Danzhou City, Hainan Province. These samples were stored in 70% ethanol, brought back to the laboratory in an ice box and stored at -80°C.

[0087] 2) A total of 365 samples of Xinong Sanen dairy goat (XNSN) were collected from the Saaneng dairy goat breeding farm in Qianyang County, Shaanxi Province. Blood samples from individuals in the breeding field were collected randomly, anticoagulated with ACD, brought back to the laboratory at low temperature in an ice box, and stored at -80°C.

...

Embodiment 3

[0143] Different STAT3 gene fragments of embodiment 3, DdeI, RsaI and TaqI digestion digestion PCR amplification

[0144] 1. DdeI, RsaI and TaqI digestion reaction digestion system

[0145]DdeI, RsaI and TaqI digestion reaction systems are all 10 μL system, including 4.0 μL of PCR product, 0.2 μL of endonuclease, 1.0 μL of 10× buffer, 1.0 μL of 0.1% BSA and 3.8 μL of deionized water.

[0146] 2. Enzyme digestion conditions:

[0147] DdeI, RsaI enzyme digestion conditions are 37 ℃ overnight (12 ~ 16h), TaqI enzyme digestion conditions are placed in a constant temperature water bath at 65 ℃ overnight (12 ~ 16h).

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Abstract

The invention discloses a method for detecting single nucleotide polymorphism of a goat signal transducer and activator of transcription (STAT3) gene and an application of the STAT3 gene. The method comprises the following steps: with a to-be-detected goat whole genome deoxyribonucleic acid (DNA) containing the STAT3 gene as a template and a primer pair P1, P2 or P3 as a primer, carrying out polymerase chain reaction (PCR) amplification on the goat STAT3 gene; digesting a PCR amplification product by using restriction enzyme DdeI, and then carrying out agarose gel electrophoresis on a digested amplification segment; and identifying the single nucleotide polymorphism of the 45204th site, the 62058th site or the 62230th site of the goat STAT3 gene according to the agarose gel electrophoresis result. Three sites can be used as molecular markers for improving the height of the xinong sannen diary goat, wherein one site is used as the molecular marker for improving cannon circumference of the Hainan black goat, and one site is used as the marker for improving the body length index of the Hainan black goat. Therefore, marker assisted selection (MAS) of growth characteristics of the native goats in China is facilitated, and quick building of goat populations with excellent genetic resources is also facilitated.

Description

technical field [0001] The invention belongs to the field of modern biotechnology and livestock breeding, and relates to the detection of gene single nucleotide polymorphism (SNP), in particular to a method and application for detecting the single nucleotide polymorphism of goat STAT3 gene. Background technique [0002] Gene polymorphism (polymorphism) refers to the difference in genome sequence between different species or different individuals within the same species. These differences are caused by nucleotide changes in DNA alleles in chromosomes, mainly including base substitutions, insertions, and deletions. and changes in copy number of repetitive sequences. In the practice of biomedicine and animal production, the detection and application of gene polymorphism are of great significance. For example, the study of the association between certain important functional gene polymorphisms and the susceptibility of diseases can clarify the susceptibility of the human body t...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCC12Q1/686C12Q1/6888C12Q2600/124C12Q2600/156C12Q2535/138
Inventor 潘传英贾文超蓝贤勇陈宏雷初朝徐铁山
Owner NORTHWEST A & F UNIV
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