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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 that can simultaneously detect the mRNA expression levels of four heat shock genes in human Jurkat cells. Methods RT-PCR system was established to detect four heat shock genes in cells simultaneously by gene cloning, gene recombination, in vitro transcription and RT-PCR. The system was used to detect the expression of heat shock genes in SW13 cells. Results The internal reference plasmids for mRNA detection of four heat shock genes were constructed and transcribed into i-RNA in vitro. The results showed that hsp70 and hsp90α showed typical heat-inducible expression in SW13 cells, hsp60 And hsp90βhad a higher level of constitutive expression, and showed varying degrees of heat-induced expression. Conclusion The newly constructed RT-PCR system can be used to detect the expression of four heat shock gene mRNAs in human Jurkat cells.