麋鹿血液cDNA文库的构建

为分离出MHCⅠ、Ⅱ基因,利用TRIzol 试剂从麋鹿血液组织中提取总RNA,经mRNA 纯化试剂盒纯化后,根据Stratagene 公司的cDNA 文库构建系统构建了麋鹿血液组织的cDNA 文库。初始文库滴度为1.96 ×106 pfu/ ml,总库容为1.96 ×106 个独立克隆;cDNA 插入片段平均长度约为1.0 kb;重组率为95.6 % 。文库各项指标均达标准经典cDNA 文库的基本要求。利用本实验室设计的可扩增MHCⅡ类DQA 基因第二外显子的引物检测扩增后的文库,得到了特异性非常好的目标条带,说明本文所建文库可以用于麋鹿MHC 相关基因的分离。

Construction of a cDNA library of blood of Elaphurus davidianus

Total RNA was extracted from the blood of Père David's deer (Elaphurus avidianus)with Trizol reagent and mRNA was further purified using a purifying kit for mRNA. According to the manual of cDNA Synthesis system from Stratagene, we constructed a cDNA library of the blood of Père David's deer. Our aim was to isolate some genes of major histocompatibility complex (MHC) for the deer. The cDNA library has a capacity of 1.96 ×106 clones, and 95.6 % of them are recombinant clones. The average size of the inserted cDNA fragments is about 1.0 kb,which meets the requirement of a standard cDNA library. The second exon of DQA gene (193 bp) was successfully amplified from the library using a primer pair designed by our lab. These results show that the blood cDNA library constructed is useful for isolation of MHCgenes of Père David's deer.