【摘要】 目的构建沉默人p38MAPK基因表达的RNA干扰的逆转录病毒表达载体。方法查找人p38MAPK cDNA序列,利用软件设计并合成能够形成互补双链并能转录出靶向人p38MAPK基因的短发夹状RNA的两条寡核苷酸序列,退火后连接至线性化载体pSIREN上,获得重组质粒pSIREN-p38MAPK,酶切和测序进行鉴定。结果双酶切重组质粒获得特定的酶切图谱,证实所合成核酸片段的正确重组;测序结果显示重组核酸片段序列与设计的序列完全相同。结论成功构建了沉默人p38MAPK基因的重组质粒pSIREN-p38MAPK。更多还原

【Abstract】 Objective It is to approach the method of constructing a RNA interfering retrovirus vector for making silence the expression of p38MAPK gene.Methods Search for the sequence of human p38MAPK cDNA,design and synthesize with software two short hairpin-shape oligonucleotide sequence that can form a complementary double-strain and transcript target human p38MARK gene,after annealing,connecting to the linear vector of pSIREN,harvesting a recombinant plasmid pSIREN-p38MAPK,identification through enzyme digestion and sequencing was made.Results Specific restriction enzyme map was obtained from double enzyme digestion,verified correctness of the recombination of synthesized nucleate fragment;sequencing showed identity between the recombinant nucleate fragment and the designed sequence.Conclusion The recombinant plasmid pSIREN-p38MAPK that can make silence the human p38MAPK gene is successfully constructed.更多还原