高原鼠兔低氧诱导因子-2α基因的克隆  

GENETIC CLONING OF HYPOXIA INDUCIBLE FACTOR-2α IN HIGH ALTITUDE HYPOXIC ADAPTATION SPECIES PLATEAU PIKA (OCHOTONA CURZONIAE)

作　　者：刘芳[1] 刘伟[2] 吴穹[1] 严云飞[1] 侯成荣[1] 

机构地区：[1]青海大学医学院 [2]青海省人民医院

出　　处：《青海医学院学报》2011年第3期151-154,158,共5页Journal of Qinghai Medical College

基　　金：青海大学中青年科研基金项目(编号:2008-QY-12)

摘　　要：目的克隆高原鼠兔(Ochotona curzoniae)低氧诱导因子-2α(Hypoxia inducible fac-tor-2,HIF-2α)基因的cDNA部分片段,以获得HIF-2αcDNA的全长序列和进行HIF-2α的表达研究。方法从高原鼠兔肺组织中提取总RNA,采用RT-PCR技术扩增出HIF-2α基因的cD-NA部分片段,与pGEM-T Easy载体连接,转化入感受态细胞JM109后,挑取阳性克隆,提取重组质粒,经酶切鉴定后进行序列测定。结果酶切分析和DNA序列分析表明,克隆所得片段大小为517bp,与预期一致。核苷酸序列和推测的氨基酸序列相似性比较显示,该片段与人、原牛、野猪、小鼠、褐家鼠HIF-2α核苷酸序列的同源性分别为90%、89%、88%、88%、86%;氨基酸序列与人、原牛、野猪、小鼠、褐家鼠HIF-2α氨基酸序列的同源性分别为98.2%、97.9%、98.1%、96.9%、96.4%。结论本实验首次成功获得高原鼠兔HIF-2α基因的cDNA部分片段,为进一步揭示高原鼠兔的低氧适应机制提供了事实依据。Objective In order to get the full-length cloning of HIF-2α cDNA and lay a foundation for researching expression of HIF-2α,cDNA fragments of HIF-2α have been cloned,which are from Plateau Pika（Ochotona curzoniae）.Methods Extracting total RNA from lung tissue of Plateau Pika,amplifying cDNA fragments of HIF-2α gene by RT-PCR technology and connecting with pGEM-T Easy vector,transforming into JM109 competent cell,taking positive clones,extracting recombinant plasmid,then testing samples after enzyme digestion.Results Restriction analysis and DNA sequence analysis showed that the length of cloned fragment was 517bp which is corresponded with expected size.Similarity comparison between nucleotide sequence and deduced amino acid sequence showed that the homologous of HIF-2α nucleotide with human,cow,wild boar,mouse,Norway rat are 90%,89%,88%,88%,86%,respectively;the homologous between amino acid sequence and the HIF-2α amino acid sequence from human,cow,wild boar,mouse,Norway rat were 98.2%,97.9%,98.1%,96.9%,96.4%,respectively.Conclusion Successfully getting cDNA fragments of HIF-2α gene from Plateau Pika laid a foundation for revealing the role of HIF-2α on hypoxia adaptation of Plateau Pika.

关 键 词：低氧诱导因子-2α 高原鼠兔 基因克隆 

分 类 号：Q78[生物学—分子生物学]