喉癌细胞酵母双杂交cDNA文库的构建

Construction of Yeast Two-hybrid cDNA Library for Hep-2 Cells

目的构建喉癌(Hep-2)细胞的酵母双杂交cDNA文库,为筛选包涵体膜蛋白分子的作用配体提供前期工作基础.方法从Hep-2细胞中提取总RNA,应用SMARTTM技术,构建以pGADT7-Rec为载体的Hep-2细胞酵母GAL4AD融合cDNA文库.结果所构建的文库展现出良好的多态性.结论可用于酵母双杂交系统的筛选.

Objective To construct yeast two-hybrid cDNA library for Hep-2 cells to screen GAL4 fusion,and provide foundation for screening ligands of Chlamydia Inclusion membrane proteins.Methods The yeast GAL4 fusion cDNA library from Hep-2 cells was constructed.Total RNA of Hep-2 cells was purified by Trizol reagents,and cDNAs of the mRNAs contained in purified total RNA sample were synthesized by SMARTTM technology,which were subsequently cloned into plasmid pGADT7-Rec.Results The library showed excellent polymorphism.Conclusion The constructed librar y could be used for yeast two-hybrid system.