论文部分内容阅读
为从基因水平上揭示正加速度 (+Gz)高耐力产生机理及寻找 +Gz高耐力相关功能性蛋白 ,利用抑制消减杂交技术分离 +Gz高耐力相关基因 .雄性SD大鼠在离心机上处理后 ,选取耐受终点在高、低两个极端的动物 ,立即取全脑 ,分离mRNA .以高耐力者为Tester ,低耐力者为Driver,利用抑制消减杂交技术进行 +Gz耐力处于高、低两个极端动物脑组织间基因表达差异显示 ,获得 +Gz高耐力大鼠脑组织相关cDNA .以高、低耐力大鼠脑组织mRNA来源的cDNA为探针 ,对获得的cDNA克隆进行斑点杂交 .分别以杂交筛选出的阳性克隆为探针 ,对高、低耐力大鼠脑组织总RNA进行Northern杂交分析 .两次杂交结果均选择高耐力组杂交信号是低耐力组 3倍以上的cDNA克隆 .经过斑点杂交筛选 ,从大鼠脑组织中获得了 6 7个在 +Gz高耐力大鼠脑组织中上调表达的cDNA克隆 .Northern杂交分析发现 ,钙离子 钙调蛋白依赖性蛋白激酶Ⅱβ亚基 (Camk2b)和一未知基因在 +Gz高耐力大鼠脑组织中的表达量增加 .结果提示 ,+Gz耐力处于高、低两个极端的大鼠脑组织基因表达有明显差异 ,这些差异表达的基因很可能与 +Gz高耐力的产生有关 ,且钙离子 钙调蛋白依赖性蛋白激酶Ⅱβ亚基和一未知基因是初步获得的与 +Gz高耐力的产生特异相关的基因
In order to reveal the mechanism of high endurance (+ Gz) tolerance and to search for functional genes related to + Gz high tolerance at the gene level, the + Gz endurance-related genes were isolated by suppression subtractive hybridization.After the male SD rats were treated on a centrifuge, Select the endpoints of the high end and low end of the two animals, immediately take the whole brain, mRNA isolation.To high endurance Tester, low endurance Driver, using suppression subtractive hybridization + Gz endurance in high and low two The difference of gene expression between the brain tissues of extreme animals showed that the cDNA of brain tissues related to + Gz high endurance rats was obtained.The dot blot of the obtained cDNA clones was carried out with the cDNAs of mRNAs from brain tissue of high and low endurance rats as The positive clones screened by hybridization were used as probes to analyze the total RNA of brain tissues of high and low endurance rats by Northern hybridization.The hybridization signals of high tolerance group and low endurance group were more than 3 times of the cDNA clone of the low endurance group Hybrid screening, we obtained 67 cDNA clones that were up-regulated in brain tissue of + Gz high-tolerance rats from rat brain.Northern hybridization showed that calcium calmodulin-dependent The expression of Camk2b and an unknown gene in the brain tissue of + Gz high endurance rats increased.The results suggest that the gene expression of + Gz endurance in the high and low end of the two extreme rat brain tissues is obvious These differences in expression of genes is likely to be related to the production of + Gz high endurance, and calcium calmodulin-dependent protein kinase Ⅱ β subunit and an unknown gene is preliminarily obtained and + Gz high endurance gene specifically related genes