A set of SNP molecular markers for screening and/or detecting ejaculatory volume and fresh sperm motility of breeding bulls
A technology for sperm motility and bulls, which is applied in the direction of recombinant DNA technology, microbial measurement/inspection, DNA/RNA fragments, etc., can solve the problems of ejaculation volume and fresh semen vitality of breeding bulls, so as to increase the economic benefits of pastures and is widely used , high accuracy effect
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Embodiment 1
[0038] The SNP locus of embodiment 1 bovine INCENP gene and the identification of genotype combination
[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 bulls used in the present invention are selected from three domestic bull breeding stations: Beijing Dairy Cow Center (83 heads), Shanghai Guangming Holstein Animal Husbandry Co., Ltd. (56 heads), and Shandong Oakes Bull Breeding Station (125 heads). Frozen bull semen was stored in liquid nitrogen for later use.
[0042] 1.2 Extraction of genomic DNA from frozen semen
[0043] 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.
[0044] 1.3 Design of primers
[0045] According to the bovine INCENP gene seq...
Embodiment 2
[0063] Example 2 Association Analysis of Cattle INCENP Gene Haplotype Combination and Bull Sperm Motility
[0064] SAS 9.0 was used to compare the correlation of different genotype combinations of INCENP gene with ejaculate volume and fresh semen motility. Its model is:
[0065] Y ijk =μ+G i +Y j +H k +e ijk
[0066] Y ijk is the observed value of ejaculation volume or sperm motility traits of bulls; 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.
[0067] The results of haplotype combination association showed that among the eight genotype combinations, the ejaculation volume of H3H4 genotype bulls was significantly higher than that of H1H4 genotype bulls (P<0.05), and was extremely significantly higher than that of H1H3, H2H4, H3H3 genotype individuals Bulls (P<0.01); H3H4 genotype combination bulls had significantly h...
Embodiment 3
[0072] Embodiment 3 detection kit
[0073] As described in Example 1, A>G at the 19970th position and T>G at the 34378th position of the bovine INCENP gene are closely related to the ejaculatory volume and fresh semen vigor of the bull. 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 according to these two SNP sites for amplification and detection.
[0074] Prepare a test kit for the detection of bull sperm motility, and the detailed components are shown in Table 2:
[0075] Table 2 The components of the test kits for the detection of bull sperm motility
[0076]
[0077] 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 components in the above kit were used to perform PCR amplificatio...
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