中文摘要：

【目的】探索热休克蛋白70-2（heat shock 70kD protein-2,HSPA2）在牦牛不同组织器官中的表达差异。【方法】选取3头1岁龄的健康青海高原雄性牦牛为研究对象,在正常生理条件下,于2013年9月中旬采集不同组织器官（心脏、肝脏、脾脏、肺脏、肾脏、脑和睾丸）样本。（1）从牦牛不同组织器官中提取RNA,将RNA反转录成第一链cDNA,参照牦牛HSPA2和β-actin基因序列（登录号为KC790105.1和DQ838049.1）设计特异性引物,首先采用RT-PCR来验证实时荧光定量PCR（RT-qPCR）是否能应用于牦牛不同组织器官中HSPA2表达差异的测定;然后采用RT-qPCR测定牦牛不同组织器官中HSPA2相对表达量的差异。（2）将牦牛不同组织器官样本4%多聚甲醛固定,制成石蜡切片,免疫组织化学法测定HSPA2在不同组织器官中的分布。采用Image-Pro Plus 6.0软件进行免疫组化图像分析,测定HSPA2阳性反应物的积分光密度,进行吸光度分析;通过SPSS 19.0统计软件,用单因素方差分析进行差异显著性测定。【结果】（1）RT-PCR结果表明RT-qPCR法可用于牦牛不同组织器官中HSPA2表达差异的测定。实时荧光定量PCR结果表明,HSPA2在睾丸中的相对表达量,分别是在脑、肾脏、心脏、脾脏、肺脏和肝脏中的83.33、97.09、111.11、133.33、222.22和285.71倍。（2）免疫组织化学结果显示,牦牛睾丸、肾脏、脑、心脏、肺脏、肝脏和脾脏中均有HSPA2的阳性表达。其中,在牦牛肾脏皮质肾小管、髓质肾小管,大脑皮质海马CA1区、小脑皮质,心肌细胞,肺泡上皮细胞,肝细胞,脾脏边缘区和红髓中有HSPA2阳性反应,着色深浅不等,大部分阳性反应位于细胞质,细胞核阳性反应极少;而在睾丸曲精小管中生精细胞细胞质和细胞核中均有HSPA2阳性反应,着色深浅不等;阴性对照组未观察到阳性反应。根据积分光密度值比较得出,HSPA2蛋白的阳性表达量在睾丸中最高,脑、肾?

英文摘要：

【Objective】 This experiment was conducted to study the differences in expression of heat shock protein 70-2（HSPA2） in different tissues and organs of yak. 【Method】 Three 1-year-old Qinghai Plateau male yaks were used in the present study. Under normal physiological conditions, the tissues and organs were obtained from healthy yak in mid-september 2013（heart, liver, spleen, lung, kidney, brain and testis）. The RNA were extracted from different tissues and organs of yak, and the RNA were reverse-transcribed into first-strand cDNA. The primers were designed specifically according to the Bos grunniens HSPA2 and β-actin gene sequences（HSPA2：KC790105.1, β-actin： DQ838049.1）. Firstly, RT-PCR were used to verify the real-time quantitative PCR（RT-qPCR） could be applied to the determination of the differences in expression of HSPA2 gene in different tissues and organs of yak. Then, real time quantitative PCR was applied to analyze the expression of HSPA2 gene. Paraformaldehyde was used to fix different tissues and organs of yak, and then made into paraffin sections. The protein localization of HSPA2 was measured by immunohistochemistry. The Image-Pro Plus 6.0 software was used to analyze immunohistochemistry, the integrated optical density of HSPA2 positive reaction was measured, and absorbance was analyzed. The SPSS19.0 software was used for significance of difference analysis. 【Result】 RT-PCR results showed that RT-qPCR can be applied to the determination of differences in expression of HSPA2 gene in different tissues and organs of yak. The results of RT-qPCR showed that comparative expression quantity of gene HSPA2 in testis was 83.33 fold than that in brain, 97.09 fold in kidney, 111.11 fold in heart,133.33 fold in spleen, 222.22 fold in lung and 285.71 fold in liver, respectively. Immunohistochemistry showed that HSPA2 expressed in the testis, kidney, brain, heart, lung, liver and spleen of yak. HSPA2 positive reaction was observed in yak renal cortical tubular, medullary tubules, cor