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Molecular marker related to lambing number character of sheep and application thereof

A molecular marker, litter size technology, applied in the determination/inspection of microorganisms, DNA/RNA fragments, recombinant DNA technology, etc., can solve the problems of slow progress in genetic improvement and low heritability, achieve good technical means and reduce costs , the effect of broad application prospects

Active Publication Date: 2014-09-17
甘肃兰天同和农业有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Reproductive traits are threshold traits, which are traits with low heritability (average 0.1), and the litter size is also affected to a certain extent under different environments and different feeding conditions. Understanding of genetic mechanisms of reproductive traits to facilitate genetic improvement of reproductive traits in sheep

Method used

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  • Molecular marker related to lambing number character of sheep and application thereof
  • Molecular marker related to lambing number character of sheep and application thereof
  • Molecular marker related to lambing number character of sheep and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0018] Cloning of embodiment 1, FSHR gene part 5'-UTR sequence

[0019] (1) Primer design

[0020] According to the sheep FSHR gene sequence (GenBank accession number: NC_019460.1), use Primer5.0 software to design upstream and downstream primers M-F and M-R, the sequence is as follows

[0021] FHSR: M-F: 5'-CGTATCTTTCCACGCCCTCT-3',

[0022] M-R: 5'-CCATCCACCCGATTGCTT-3'.

[0023] (2) Cloning and sequencing of PCR products

[0024] Ligate the purified PCR product with the pMD-18 T vector (purchased from Takara) in a water bath at 4°C overnight; take 100-120 μl of competent cells in a 1.5 ml Ependorff tube under sterile conditions, and add 5 μl of the ligation product to Mix well, place on ice for 30 min, heat shock at 42°C for 90 s, then ice-bath for 3-4 min, add 400 μl LB liquid medium without antibiotics, and incubate at 37°C for 45 min with shaking. Take 100 μl and spread it on the agar plate of isopropylthio-β-D-galactoside (IPTG) X-gal, place it flat at...

Embodiment 2

[0027] Embodiment 2, establishment of PCR-RFLP diagnostic method

[0028] (1), primer sequence

[0029] FSHR: M-F: 5'-CGTATCTTTCCACGCCCTCT-3',

[0030] M-R: 5'-CCATCCACCCGATTGCTT-3'.

[0031] (2) PCR amplification conditions

[0032] The total volume of the PCR reaction is 20 μl, including about 100 ng of sheep genomic DNA, containing 1× buffer (purchased from Promega), 1.5 mmol / L MgCl 2 , the final concentration of dNTP is 150 μmol / L, the final concentration of primer is 0.4 μmol / L, 2U Taq DNA polymerase (Promega). The PCR amplification program was: 94°C for 3 min, 35 cycles of 94°C for 30 s, 58°C for 30 s, then 72°C for 25 s, and finally 72°C for 5 min. PCR reaction products were detected by 2% agarose gel electrophoresis. The result is as figure 1 shown. A 244bp specific fragment was amplified, which was located in the 5'-UTR. As a result of sequencing, it was found that there was a BsiEI restriction site (CGRY↓CG) in the 244bp fragment, and the 90th ...

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Abstract

The invention belongs to the technical field of preparation of molecular markers for livestock, and particularly relates to a preparation method and application of a molecular marker which is related to lambing number character of sheep and used for sheep marker assisted selection. The molecular marker is obtained by FSHR gene cloning, and the nucleotide sequence of the molecular marker is as shown in a sequence table SEQ ID NO:1. One T88 - C88 base substitution is generated at the 88 bp of the sequence table SEQ ID NO:1, thus resulting in BsiEI-RFLP digestion polymorphism. The invention also discloses a primer for amplifying a part of DNA (deoxyribonucleic acid) sequence of an FSHR gene and a method used for polymorphism detection, and provides a new molecular marker for selection assisted by the marker related to lambing number character of sheep.

Description

technical field [0001] The invention belongs to the technical field of livestock molecular marker preparation, and in particular relates to a preparation method and application of a molecular marker related to sheep litter size traits used as sheep marker-assisted selection. Background technique [0002] According to the statistics of FAO (2012), there are 187 million sheep in my country, accounting for 16% of the world's breeding (1.169 billion), ranking first in the world (http: / / www.fao.org / home / en / ), It is the largest producer and consumer of mutton. The population genetic improvement of sheep is of great significance to ensure the supply of mutton, promote the development of the national economy and improve people's living standards. The reproductive traits of sheep, especially the lamb size is one of the most important economic traits in modern sheep production. With the implementation of the national ecological protection strategy, the space for grazing and raising s...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/11C12Q1/68
Inventor 李发弟王维民翁秀秀刘世佳潘香羽马友记唐德富李冲
Owner 甘肃兰天同和农业有限公司
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