摘要
[目的]克隆人Notch信号通路中配体Jagged1基因并进行真核表达。[方法]采用RT-PCR的方法从Hela细胞总RNA中获取Jagged1基因,并克隆至携带FLAG标签的真核表达载体pCMV-Tag4。经酶切、PCR和测序鉴定后,将重组质粒Jagged1-pCMV-Tag4瞬时转染HEK 293T细胞,通过Q-PCR和Western blot检测目的蛋白的表达。[结果]成功构建了真核表达质粒Jagged1-pCMV-Tag4并在HEK 293T细胞中瞬时表达。[结论]Jagged1在HEK 293T细胞中实现瞬时表达,为稳定表达和进一步研究Jagged1/Notch信号通路奠定基础。
[Objective]To clone and express human Notch ligand Jaggedl in eukaryotic cells. [ Methods] Jaggedl gene was amplified from total RNA in Hela cells by RT - PCR, and cloned into eukaryotic expression vector pCMV - Tag4 containing FLAG to construct Jaggedl - pCMV - Tag4. The recombinant plasmid was transiently transfected into HEK 293T cells. Real - time quantitative PCR and Western blot were performed for further confirmation. [ Results ] The eukaryotic expression plasmid Jaggedl -pCMV- Tag4 was constructed successfully and was transfected into HEK 293T cell. [ Conclusion] Jaggedl is ex- pressed in HEK 293T cell which provides a foundation for further investigation of Jaggedl/Notch signaling pathway.
出处
《生物技术》
CAS
CSCD
北大核心
2015年第5期437-441,共5页
Biotechnology
基金
国家高技术研究发展技术(863计划)("蛋白质测序新技术新装备及配套试剂国产化"
No.2014AA020909)
国家自然科学基金-广东省联合基金重点项目("番荔枝内酯防治肿瘤的关键科学问题及纳米靶向制剂成药性研究"
No.U1401223)
国家自然科学基金面上项目("幽门螺杆菌Lpp20抗原引起的免疫优势CD4+T细胞应答及与胃部疾病的相关性"
No.81470831)
广东省科技计划创新载体建设项目("珠海南医大生物医药公共服务平台"
No.2013B090800036)资助
