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SNP (single-nucleotide polymorphism) sites related to thawed stud bull frozen semen activity and deformity rate and application thereof

A technology of bulls and frozen semen, applied in the direction of recombinant DNA technology, microbial measurement/testing, biochemical equipment and methods, etc., can solve the problems of different related traits, achieve low cost, reduce blindness, and reduce costs

Pending Publication Date: 2016-08-24
DAIRY CATTLE RES CENT SHANDONG ACADEMY OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are as many as 2,246 known SNP sites in the ARID4A gene in Bos Taurus, and the associated traits of different SNP sites are often different. SNP molecular marker is the main problem faced by the technology in the art
In the prior art, there is no report on the SNPs of the ARID4A gene and its genotype combination that affect the vigor and deformity rate of the bull's frozen semen after thawing.

Method used

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  • SNP (single-nucleotide polymorphism) sites related to thawed stud bull frozen semen activity and deformity rate and application thereof
  • SNP (single-nucleotide polymorphism) sites related to thawed stud bull frozen semen activity and deformity rate and application thereof
  • SNP (single-nucleotide polymorphism) sites related to thawed stud bull frozen semen activity and deformity rate and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Example 1: Identification of SNP sites and genotype combinations of bovine ARID4A gene

[0048] The invention adopts the means of direct sequencing, CRS-PCR and PCR-RFLP to identify the SNP of bovine ARID4A.

[0049] 1.1 Experimental materials

[0050] The bulls used in the present invention are selected from two domestic bull breeding stations: Beijing Dairy Cow Center (85 heads) and Shandong Oakes Breeding Bull Station (130 heads). Frozen bull semen was stored in liquid nitrogen for later use.

[0051] 1.2 Extraction of genomic DNA from frozen semen

[0052] The high-salt method was used to extract the genomic DNA from the frozen semen, and the purity and concentration of the DNA samples were detected by a nucleic acid and protein analyzer, and finally diluted to 50 ng / μl, and stored at 4°C for later use.

[0053] 1.3 Design of primers

[0054] According to the bovine ARID4A gene sequence (accession number: AC_000167.1) provided by NCBI, utilize Primer Premier5.0 ...

Embodiment 2

[0083] Example 2: Association analysis of bovine ARID4A gene haplotype combinations and bull sperm motility

[0084] SAS 9.0 was used to compare the correlation between different genotype combinations of ARID4A gene, deformity rate and frozen semen viability after thawing. Its model is:

[0085] Y ijk =μ+G i +Y j +H k +e ijk

[0086] Y ijk is the observed value of bull deformity rate or sperm motility traits; μ is the average value of the population; G i : fixed effect of haplotype combination; Y j : fixed effect of season; H k : fixed effect of the number of sessions; e ijk : random residual effect.

[0087] The results of haplotype combination association showed that among the six genotype combinations, the deformity rate of H4H4 genotype bulls was significantly lower than that of H1H1 and H1H3 genotype bulls (P<0.05); The post vigor was significantly higher than that of H1H3 (P<0.05) genotype individual bulls (Table 1).

[0088] Table 1 Correlation analysis bet...

Embodiment 3

[0093] Embodiment 3 detection kit

[0094] As described in Example 1, G>T at position 53 and G>A at position 826 of the bovine ARID4A gene are closely related to the deformity rate of breeding bulls and the vigor after thawing of frozen semen. Therefore, it is possible to invent a kind of ARID4A gene SNP molecular marker detection kit suitable for screening the deformity rate of breeding bulls and the viability of frozen semen after thawing, that is, according to these two SNP sites, the specific primers of bovine ARID4A gene are designed to amplify and detection.

[0095] Prepare a test kit for the detection of bull sperm motility, and the detailed components are shown in Table 2:

[0096] Table 2 The components of the test kits for the detection of bull sperm motility

[0097]

[0098] We first collected the semen of the breeding bulls, stored them in liquid nitrogen for later use, and extracted the total DNA from the frozen semen using the high-salt method. The compon...

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Abstract

The invention relates to a set of SNP (single-nucleotide polymorphism) molecular markers for screening and detecting stud bull deformity rate and thawed frozen semen activity. SNP sites are positioned on 53rd and 826th bases of ARID4A gene; the two SNP sites have multiple genotype combinations in stud bull populations; and the genome DNA of the stud bull frozen semen is extracted to detect the genotype combination of the ARID4A gene and predict the stud bull deformity rate and thawed frozen semen activity. The invention successfully screens out the SNPs of the stud bull ARID4A gene, illuminates the relevance of the stud bull ARID4A gene mononucleotide polymorphism to the stud bull deformity rate and thawed frozen semen activity, provides a simple and efficient method for detecting the stud bull deformity rate and thawed frozen semen activity, and develops the corresponding detection kit. The kit has the advantages of low cost, high reliability and stability, high efficiency and the like, and can be used for screening early high-quality stud bulls.

Description

technical field [0001] The invention belongs to the technical field of livestock molecular biology, and specifically relates to a group of SNP sites related to the vigor and deformity rate of breeding bulls after thawing from frozen semen and their application, especially relates to the detection of the relationship between the deformity rate and frozen semen of breeding bulls by molecular marker technology. The SNPs of the ARID4A gene related to the vigor level after thawing of semen provide a method and a kit for detecting the deformity rate of breeding bulls and the vigor level after thawing of semen, and are 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. Today, with artificial insemination and frozen semen technologies being widely p...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6888C12Q2600/124C12Q2600/156
Inventor 孙艳刘娟李秋玲王长法杨春红姜强王秀革鞠志花张燕黄金明仲跻峰
Owner DAIRY CATTLE RES CENT SHANDONG ACADEMY OF AGRI SCI
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