奶牛HSP70-A8基因多态、表达及其与DHI性能和生理指标关联分析

发布时间：2018-01-20 16:33

本文关键词： 奶牛 HSP70-A8 遗传多态性表达水平 DHI性能生理指标　出处：《安徽农业大学》2015年硕士论文　论文类型：学位论文

【摘要】：热应激蛋白70(HSP70)作为最重要的分子伴侣参与到动物机体内的热应激反应之中,为了研究HSP70-A8基因对奶牛生产性能和耐热性能的影响,以397头荷斯坦奶牛为主要研究对象,采用单链构象多态性(PCR-SSCP)技术,对HSP70-A8基因编码区(664 to 5089)进行克隆与多态性研究,并分析其与奶牛DHI生产性能的相关性；以10头荷斯坦牛和娟×荷杂交牛为研究对象,采用实时荧光定量技术(RT-PCR)分析和比较了不同遗传型奶牛在不同季节(环境温度)下HSP70-A8基因在血液组织中的相对表达水平,同时分析了表达水平与生理指标的关系。取得的结果如下：1.荷斯坦奶牛HSP70-A8基因的多态性及其遗传分析利用PCR-SSCP方法进行检测,在HSP70-A8基因的第2-8外显子上均没有检测到突变,在第1外显子1522BP处存在G→C的错义突变,使编码的氨基酸由谷氨酰胺变成组氨酸。检测到有三种基因型AA、AB和BB。其中AA基因型频率为38.56%,AB基因型频率分别为31.03%,BB基因型频率分别为30.41%；多态信息含量(PIC)为0.3733,在该位点均处于中度多态；卡方检验表明试验牛群体处于哈迪-温伯格不平衡状态。2.荷斯坦奶牛HSP70-A8基因的多态性与生产性能的关联分析对该位点的不同基因型与生产性能进行关联分析,结果表明：AA型和AB型个体的305d标准乳产量、总经济校正奶量、305d乳脂产量、305d乳蛋白产量和总乳蛋白产量极显著低于BB型个体(P0.01)；AA型个体的标准乳总产量、总产奶量、305d经济校正奶量和总乳脂产量显著低于AB型个体(P0.05),AB型又极显著低于BB型个体(JP0.01);AA型个体的体细胞数极显著低于AB型个体和BB型个体(P0.01)。3.不同遗传型奶牛在不同环境温度下HSP70-A8基因表达水平的比较分析采用实时荧光定量PCR方法分析HSP70-A8基因在不同季节中的相对表达量,结果表明：夏季HSP70-A8基因在荷斯坦牛和F1代中的表达量极显著低于在F2代中的表达量(P0.01),秋季和冬季HSP70-A8基因在三个品种中的表达差异不显著；荷斯坦奶牛HSP70-A8基因在三个季节中的表达量差异不显著,F1代HSP70-A8基因在三个季节的表达量差异不显著,F2代HSP70-A8基因在夏季的表达量显著高于冬季(P0.01)。4.奶牛主要生理指标差异性分析及其与HSP70-A8基因表达水平关联分析对HSP70-A8基因表达量和生理指标进行分析,结果表明：奶牛在夏季的呼吸频率显著高于秋冬季节(P0.01),夏季的直肠温度明显高于冬季(P0.01),奶牛在三个季节中的心跳次数差异不显著。在三个季节中,不同品种之间的呼吸频率、直肠温度和心跳次数差异均不显著。HSP70-A8的表达量和奶牛的呼吸频率、心跳次数和直肠温度均无明显相关性。研究结果提示HSP70-A8基因对于奶牛的生产性能具有显著性影响,可以作为奶牛生产性能选择的候选遗传分子标记,BB基因型个体可以作为生产性状优势个体进行培育。
[Abstract]:Heat stress protein 70 (HSP70), as the most important molecular chaperone, is involved in the heat stress reaction in animal body. In order to study the effect of HSP70-A8 gene on the performance and heat resistance of dairy cattle, single strand conformation polymorphism (SSCP) technique was used to study 397 Holstein cows. Cloning and polymorphism of HSP70-A8 gene coding region from 664 to 5089) were studied, and the correlation between DHI production performance and DHI was analyzed. Ten Holstein cattle and Juan 脳 Hollow hybrid cattle were studied. The relative expression level of HSP70-A8 gene in blood tissues of dairy cattle with different genetic types in different seasons (ambient temperature) was analyzed and compared by real-time fluorescence quantitative technique RT-PCR. At the same time, the relationship between expression level and physiological indexes was analyzed. The results were as follows: 1. The polymorphism of HSP70-A8 gene in Holstein cows and its genetic analysis were detected by PCR-SSCP method. No mutation was detected in exon 2-8 of HSP70-A8 gene, and G existed at exon 1522BP of exon 1. 鈫扵he missense mutation of C resulted in the transformation of the encoded amino acid from glutamine to histidine. Three genotypes were detected, AAAAB and BB.The frequency of AA genotype was 38.56%. The frequency of AB genotype was 31.03and BB genotype was 30.41; The polymorphic information content (Pi) was 0.3733, which was moderately polymorphic at this locus. The chi-square test showed that the population of the tested cattle was in the Hady-Weinberg disequilibrium state. 2. Correlation analysis of the polymorphism of HSP70-A8 gene and the production performance of Holstein dairy cattle with different genotypes and productivity of the locus. To be able to perform correlation analysis. The results showed that the 305d standard milk yield and the total corrected milk yield were 305d and 305d respectively. The milk protein yield and total milk protein yield at 305d were significantly lower than those of BB type individuals (P 0.01). The total standard milk yield, total milk yield and total milk fat yield of AA individuals were significantly lower than those of AB individuals (P 0.05). AB type was significantly lower than BB type individuals. The number of somatic cells in AA individuals was significantly lower than that in AB individuals and BB individuals (P 0.01). Comparative analysis of HSP70-A8 gene expression levels in different genotypes of dairy cattle under different ambient temperature using real-time fluorescence quantitative PCR method to analyze the phase of HSP70-A8 gene in different seasons. Yes, expression. The results showed that the expression of HSP70-A8 gene in Holstein cattle and F1 generation in summer was significantly lower than that in F2 generation (P 0.01). There was no significant difference in the expression of HSP70-A8 gene between autumn and winter. There was no significant difference in the expression of HSP70-A8 gene among the three seasons in Holstein dairy cattle. There was no significant difference in the expression of HSP70-A8 gene in F1 generation in the three seasons. The expression of HSP70-A8 gene in F2 generation in summer was significantly higher than that in winter (P0.01). Analysis of the difference of the main physiological indexes and the correlation analysis between the expression level of HSP70-A8 gene and the expression level of HSP70-A8 gene in dairy cattle. The expression quantity and physiological index of HSP70-A8 gene were analyzed. The results showed that the respiratory frequency in summer was significantly higher than that in autumn and winter, and the rectal temperature in summer was significantly higher than that in winter. There was no significant difference in the number of heart beats in the three seasons. In the three seasons, the respiration rate among different breeds was not significant. There was no significant difference in rectal temperature and heart rate. The expression of HSP70-A8 and the respiratory rate of dairy cattle were not significant. There was no significant correlation between heart rate and rectal temperature. The results showed that HSP70-A8 gene had a significant effect on the performance of dairy cattle. The individuals of BB genotype which can be used as candidate genetic markers for performance selection of dairy cattle can be bred as dominant individuals of production traits.
【学位授予单位】：安徽农业大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S823

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