摘要
目的 构建人N端截断型脂多糖结合蛋白 (tLBP)基因的杆状病毒表达载体 ,为后续的蛋白表达及其抗内毒素保护作用研究奠定基础。方法 采用PCR技术从人全长LBP基因cDNA中扩增长度为 6 95bp的基因片段 ,经pGEM T载体亚克隆筛选和序列分析后 ,应用Bac To Bac杆状病毒表达系统 ,基因重组和外源基因转座分别构建转移质粒pFASTBACtLBP和穿梭质粒pBacmidtLBP。结果 经PCR、琼脂糖凝胶电泳、酶切分析及序列测定 ,所克隆的基因片段为 70 0bp左右的tLBP ,并证实其目的基因重组载体发生了特异转座和病毒重组。结论 本实验成功克隆了tLBP目的基因片段并构建其杆状病毒表达载体。
Objective To construct a baculovirus expression vector of human truncated lipopolysaccharide binding protein for further study on protein expression and anti endotoxin protection. Methods A fragment of 695 bp from the amino terminal of human LBP cDNA was cloned by using PCR. Plasmid pFASTBACtLBP and pBacmidtLBP were constructed with Bac to Bac baculovirus expression system respectively following subclone screening and sequencing for pGEM T. The constructed recombinant was then identifed with PCR and agarose gel electrophoresis. Results A 700±bp gene fragment of tLBP was found and specific transposition and virus recombination were observed in the constructed plasmids. Conclusion The tLBP gene fragment was cloned and constructed to baculovirus expression vectors succcessfully.
出处
《第三军医大学学报》
CAS
CSCD
北大核心
2001年第6期637-640,共4页
Journal of Third Military Medical University
基金
国家自然科学基金资助项目 (3 9970 75 7)
全军"九五"医药卫生科研基金资助项目 (98M0 91 )
