摘要
目的克隆Ang-1基因,构建Ang-1/Pshuttle/Padeasy-1腺病毒表达载体,为进一步研究Ang-1防治卵巢癌病变的血管异质性提供理论基础。方法提取流产胎儿肝脏组织总RNA,RT-PCR扩增出Ang-1基因片段,并将其克隆到Pshuttle载体中进行序列分析,经PacⅠ线性化、连接到腺病毒表达载体Padeasy-1中,将Padeasy-1包装进脂质体、转染入人胚肾293细胞,测定病毒生物活性滴度。结果从流产胎儿肝脏组织总RNA中,扩增出1.4kb的cDNA片段,成功构建了Padeasy-1/Pshuttle/Ang-1表达载体。结论从流产胎儿肝脏组织中成功克隆Ang-1基因,成功构建Padeasy-1/Pshuttle/Ang-1表达载体,Padeasy-1/Pshuttle/Ang-1表达载体的滴度为1×1011PFU/mL。
Objective To clone angiopoietin-1 genes and construct Ang-1/Pshuttle/Pageasy-1 adenovirus expression vector, so as to make theory base for the further research on the Ang-1 prevention and cure blood vessel heterogenous of ovarian caner pathological changes. Methods Extracting mRNA from abortion embryo liver,amplified Ang-1 gene by RT-PCR and cloned into Pshuttle vector and sequenced,linearized by Pac Ⅰ and connected to adenovirus expression vector Padeasy-1, packaging Padeasy into liposome,transfect to 293 cells, determination of virus biological activity titer. Results From abortion embryo liver total RNA, RT- PCR amplified 1. 4kb cDNA fragment, successfully constructing Padeasy-1/Pshuttle/Ang-1 express vector. Conclusion Cloned Ang-1 gene and successfully constructing Padeasy-1/Pshuttle/Ang-1 express vector,it＇s titer is 1 × 10^11PFU/mL.
出处
《重庆医学》
CAS
CSCD
北大核心
2009年第10期1190-1192,共3页
Chongqing medicine
基金
国家自然科学基金资助项目(30600673)
"十一五"全军科技计划青年基金项目(06Q054)。
