荷斯坦母牛TLR1基因多态性及其与体细胞评分关系

 【目的】阐明Toll-like receptor 1（TLR1）基因多态性与荷斯坦母牛乳房炎抗性之间的关系,为奶牛乳房炎抗性的标记辅助选择提供依据。【方法】采用PCR-SSCP和PCR-RFLP检测208头荷斯坦母牛TLR1基因多态性,用最小二乘法分析基因多态性与荷斯坦母牛体细胞评分（somatic cell score,SCS）的关系。【结果】TLR1基因mRNA序列中存在G1409A、A1475C、G1550A和G1596A 突变位点,其中G1596A突变使异亮氨酸变为缬氨酸;G1409A突变位点经BsiYⅠ酶切产生2种基因型AA和AB,频率分别为0.375和0.625,χ2检验表明荷斯坦母牛在该位点偏离Hardy-Weinberg平衡;AA和AB基因型SCS最小二乘均值差异不显著（P＞0.05）;A1475C突变位点经BstXⅠ酶切产生2种基因型CC和CD,频率分别为0.178和0.822,荷斯坦母牛在该位点偏离了Hardy-Weinberg平衡;CC和CD基因型SCS最小二乘均值差异不显著（P＞0.05）;G1550A突变位点经BsiYⅠ酶切产生3种基因型EE、EF和FF,频率分别为0.236、0.370和0.394,荷斯坦母牛在该位点偏离了Hardy-Weinberg平衡;EE、EF和FF基因型SCS最小二乘均值差异均不显著（P＞0.05）;G1596A突变位点经BclⅠ酶切产生3种基因型GG、GH和HH,频率分别为0.260、0.567和0.173,荷斯坦母牛在该位点处于Hardy-Weinberg平衡;HH基因型SCS最小二乘均值极显著低于GH基因型所对应的值（P＜0.01）,极显著低于GG基因型所对应的值（P＜0.0001）;GH基因型SCS最小二乘均值极显著低于GG基因型所对应的值（P＜0.01）。【结论】对奶牛乳房炎抗性,HH是最有利基因型,GG是最不利基因型,初步证明TLR1基因G1596A多态位点的H等位基因是提高奶牛乳房炎抗性的一个潜在的DNA标记。

Polymorphisms of TLR1 Gene and Their Relationship with Somatic Cell Score in Holstein Cows

【Objective】 The objectives of the present study were to elucidate the relationship between Toll-like receptor 1 (TLR1) gene and mastitis resistance of Holstein cows and to provide a scientific basis for marker-assisted selection for mastitis resistance of dairy cattle. 【Method】 Polymorphisms of all exons of TLR1 gene were detected in 208 Holstein cows by PCR-SSCP and PCR-RFLP. The relationship between polymorphisms of TLR1 gene and somatic cell score (SCS) in Holstein cows was analyzed by least squares linear model. 【Result】 Four polymorphic sites, G1409A, A1475C, G1550A and G1596A, were found, and G1596A mutation resulted in an amino acid change of isoleucine→valine. For G1409A polymorphic site, two genotypes (AA and AB) were found in 208 Holstein cows by digesting the PCR products with restriction endonuclease BsiYⅠ, and the frequency of AA and AB genotypes was 0.375 and 0.625, respectively. Chi-square test showed that Holstein cows were not in Hardy-Weinberg equilibrium. Least squares mean of SCS for AA was not significantly lower than that for AB (P＞0.05). For A1475C polymorphic site, two genotypes (CC and CD) were found in 208 Holstein cows by digesting the PCR products with restriction endonuclease BstXⅠ, and the frequency of CC and CD genotypes was 0.178 and 0.822, respectively. Holstein cows were not in Hardy-Weinberg equilibrium. Least squares mean of SCS for CC was not significantly lower than that for CD (P＞0.05). For G1550A polymorphic site, three genotypes (EE, EF and FF) were found in 208 Holstein cows by digesting the PCR products with restriction endonuclease BsiYⅠ, and the frequency of EE, EF and FF genotypes was 0.236, 0.370 and 0.394, respectively. Holstein cows were not in Hardy-Weinberg equilibrium. The difference of the least squares means for SCS between EE, EF and FF was insignificant (P＞0.05). For G1596A polymorphic site, three genotypes (GG, GH and HH) were found in 208 Holstein cows by digesting the PCR products with restriction endonuclease BclⅠ, and the frequency of GG, GH and HH genotypes was 0.260, 0.567 and 0.173, respectively. Holstein cows were in Hardy-Weinberg equilibrium. Least squares mean of SCS for HH was significantly lower than that for GH (P＜0.01) or GG (P＜0.0001), least squares mean of SCS for GH was significantly lower than that for GG (P＜0.01). 【Conclusion】 HH was the most favorable genotype, GG was the most unfavorable genotype for mastitis resistance. The results preliminarily indicated that allele H of G1596A polymorphic site of TLR1 gene is a potential DNA marker for improving mastitis resistance in dairy cattle.