Group of SNP (single nucleotide polymorphism) molecular markers for screening and detecting ejaculatory amount and fresh sperm activity level of stud bulls

A bull and sperm measurement technology, applied in the direction of recombinant DNA technology, microbial determination/inspection, DNA/RNA fragments, etc., can solve the problems of ejaculation volume and fresh sperm vitality of breeding bulls, increase the economic benefits of the pasture, reduce blindness performance, cost reduction

Active Publication Date: 2015-11-11
DAIRY CATTLE RES CENT SHANDONG ACADEMY OF AGRI SCI
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AI Technical Summary

Problems solved by technology

In the prior art, there is still no report on the SNPs of the INCENP gene and the genotype combination of th

Method used

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  • Group of SNP (single nucleotide polymorphism) molecular markers for screening and detecting ejaculatory amount and fresh sperm activity level of stud bulls
  • Group of SNP (single nucleotide polymorphism) molecular markers for screening and detecting ejaculatory amount and fresh sperm activity level of stud bulls
  • Group of SNP (single nucleotide polymorphism) molecular markers for screening and detecting ejaculatory amount and fresh sperm activity level of stud bulls

Examples

Experimental program
Comparison scheme
Effect test

Example Embodiment

[0038] Example 1 Identification of SNP site and genotype combination of bovine INCENP gene

[0039] The invention adopts the means of direct sequencing and PCR-RFLP to carry out SNP identification on bovine INCENP.

[0040] 1.1 Experimental materials

[0041] The bull used in the present invention is selected from three domestic bull breeding stations: Beijing Dairy Cow Center (83 heads), Shanghai Guangming Holstein Animal Husbandry Co., Ltd. (56 heads), and Shandong Aux Breeding Bull Station (125 heads). Bull frozen semen is stored in liquid nitrogen for later use.

[0042] 1.2 Extraction of frozen sperm genomic DNA

[0043] The frozen sperm genomic DNA was extracted by the high-salt method, and the purity and concentration of the DNA sample were detected by a nucleic acid protein analyzer, and finally diluted to 50ng / μl, and stored at 4°C for later use.

[0044] 1.3 Design of primers

[0045] According to the bovine INCENP gene sequence provided by NCBI (accession number: AC_000186.1),...

Example Embodiment

[0063] Example 2 Association analysis of bovine INCENP gene haplotype combination and bull sperm motility

[0064] Using SAS9.0, the correlation between different genotype combinations of bovine INCENP gene and ejaculation volume and fresh sperm vitality was compared. The model is:

[0065] Y ijk =μ+G i +Y j +H k +e ijk

[0066] Y ijk Is the observed value of bull's ejaculation volume or sperm motility traits; μ is the population average; G i : Fixed effect of haplotype combination; Y j : Seasonal fixed effects; H k : Fixed effect of the session; e ijk : Random residual effect.

[0067] The results of haplotype combination association showed that among the 8 genotype combinations, the ejaculate volume of bulls with H3H4 genotype was significantly higher than that of bulls with H1H4 genotype (P <0.05), extremely significantly higher than H1H3, H2H4, H3H3 genotype individual bulls (P <0.01); H3H4 genotype combination bull individual fresh sperm vitality was significantly higher ...

Example Embodiment

[0072] Example 3 Detection kit

[0073] As described in Example 1, the A> at position 19970 of the bovine INCENP gene G and the 34378th T> G is closely related to the ejaculation volume and freshness vitality of bulls. Therefore, it is possible to invent an INCENP gene SNP molecular marker detection kit suitable for screening bull ejaculation volume and fresh sperm vigor, that is, design specific primers for bovine INCENP gene for amplification and detection according to these two SNP sites.

[0074] A kit for detecting the level of sperm motility of bulls was prepared. The detailed components are shown in Table 2:

[0075] Table 2 Components of the kit for testing bull sperm motility

[0076]

[0077] We first collected the semen of the bulls and stored them in liquid nitrogen for later use. The total DNA of frozen semen was extracted by the high-salt method. The components in the above kit are used for PCR amplification using the extracted frozen sperm DNA as a template. Then, the...

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Abstract

The invention relates to a group of SNP (single nucleotide polymorphism) molecular markers for screening and detecting the ejaculatory amount and the fresh sperm activity level of stud bulls. Specifically, SNPs sites are positioned at the 19970th and 34378th basic groups of INCENP genes, the two SNPs sites have multiple types of genotype combinations in stud bull groups, and the genotype combinations of the INCENP genes of frozen sperms of the stud bulls are detected and the ejaculatory amount and the fresh sperm activity of the stud bulls are predicted by extracting the genome DNA of the frozen sperms of the stud bulls. The SNPs of the INCENP genes of the stud bulls are successfully screened out, the correlation between the mononucleotide polymorphism of the INCENP genes of the stud bulls and the ejaculatory amount and the fresh sperm activity of the stud bulls is illuminated for the first time, a simple, convenient and efficient method for detecting the ejaculatory amount and the fresh sperm activity of the stud bulls is provided, and a corresponding detection kit is developed. The kit has the advantages of low cost, stable reliability, high efficiency and the like, and can be used for screening early high-quality stud bulls.

Description

technical field [0001] The present invention relates to the technical field of livestock molecular genetics, in particular to a set of SNP molecular markers for screening and / or detecting ejaculatory volume and fresh semen vigor of breeding bulls, and in particular to detection of ejaculatory volume and fresh semen activity of breeding bulls through molecular marker technology The SNPs locus of the INCENP gene related to the level of vigor provides a method and kit for detecting the amount of ejaculation and the vigor of fresh semen of bulls, and is applied to the early breeding of excellent livestock reproductive traits. Background technique [0002] Milk has high nutritional value and is an indispensable food in our daily life. Chinese Holstein cows are the breed with the widest breeding range and the largest milk yield in China. Chinese Holstein bulls are an important part of dairy cow breeding. The quality of their semen not only determines the economic benefits of the ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6888C12Q2600/124C12Q2600/156
Inventor 王长法刘娟孙艳姜强杨春红刘保申鞠志花王秀革黄金明仲跻峰
Owner DAIRY CATTLE RES CENT SHANDONG ACADEMY OF AGRI SCI
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