检测4种热休克基因表达水平的RT-PCR体系的建立及其应用

Establishment and Application of a RT-PCR System for Quantifying mRNA of 4hsp Genes in Human Cells

目的建立能同时检测人Jurkat细胞中4种热休克基因mRNA表达水平的RT-PCR体系。方法采用基因克隆、基因重组、体外转录、RT-PCR技术,建立能同时检测细胞中4种热休克基因的RT-PCR体系,并将此体系用于检测SW13细胞中热休克基因的表达。结果构建了用于4种热休克基因mRNA检测的内参照质粒,并经体外转录成内参照RNA(i-RNA);用于SW13细胞中结果显示:hsp70、hsp90α呈典型的热诱导表达,hsp60和hsp90β具有较高水平的组成性表达,并且呈不同程度的热诱导表达。结论新构建的RT-PCR体系可以用于检测人Jurkat细胞中4种热休克基因mRNA的表达水平。

Objective To establish a RT-PCR system for detecting mRNA expression level of4hsp genes in human cells.Methods RT-PCR system was established with gene cloning,gene recombination,in vitro transcription and RT-PCR techniques;The detection of expression level of4hsp genes in SW13cell was carried out with this system.Results In SW13cell,hsp70and hsp90αwere typical heat shock induced genes,while hsp60and hsp90βwere efficiently expressed and further induced by heat-shock to various extent.Conclusions In our hands novel RT-PCR system can be used to detect mRNA expression level of4human hsp genes.