摘要
目的构建小鼠PICK1基因的真核表达载体myc-PICK1。方法从小鼠脑组织中提取总RNA,RT-PCR扩增PICK1序列,所得片段及真核表达载体p RK5-myc用Sal I和Not I双酶切,后连入p RK5-myc载体中。重组质粒经Sal I和Not I双酶切鉴定后送公司测序。结果 PCR扩增得到预期1.2 kb大小的目的片段,经Sal I和Not I双酶切鉴定正确重组质粒送公司测序,结果与预期一致。myc-PICK1重组质粒转染Hela细胞,进行免疫染色能看到定位于胞浆的蛋白表达。结论成功构建了myc-PICK1载体,为后续研究奠定了基础。
Objective In order to construct Eukaryotic expression vector of myc-PICK1. Methods RNA of mice brain was extracted, PICK1 fragments were obtained by RT-PCR and were sub-cloned into p RK5 with myc tag to get myc-PICK1, and then recombinants were sequenced after confirmation by digested with Sal I and Not I. Results About 1.2 kb fragments of myc-PICK1 was amplified through RT-PCR, correct recombinant plasmid was sent to the company to sequence after confirming by digested with Sal I and Not I, and the sequence was correct. Recombinant protein myc-PICK1 was observed to express in cytoplasm after being transfected to Hela cells and being immunostained. Conclusion The recombinant plasmid myc-PICK1 is successfully constructed.
出处
《解剖学研究》
CAS
2015年第4期241-244,共4页
Anatomy Research
基金
成都大学自然科学基金(2011XJZ14)
成都大学人才工程科研启动基金
关键词
PICK1
真核表达载体
小鼠
Protein that interacts with C kinase 1（PICK1）
Eukaryotic expression vector
Mice
