中文摘要：

目的：为深入研究热休克蛋白27（heat shock protein27，HSP27）的生理与病理功能，应用PAdEasy腺病毒载体系统构建带荧光蛋白的小鼠HSP27重组腺病毒，并进行鉴定。方法：采用基因技术，将目的基因克隆至穿梭质粒pAdTrack-CMV中，通过PCR、序列测定、酶切鉴定；利用PAdEasy系统进行重组，在293细胞中包装和扩增，然后体外感染小鼠精原母细胞，Western blot检测HSP27蛋白表达。结果：测序和酶切鉴定重组腺病毒质粒构建正确；经293细胞包装后可观察到绿色荧光，收获高滴度重组腺病毒体外感染小鼠精原母细胞．并且可正确表达HSP27。结论：应用PAdEasy系统重组技术成功构建了小鼠HSP27带荧光蛋白腺病毒载体，并在小鼠精母细胞系进行体外有效表达，为进一步研究HSP27的功能奠定基础。

英文摘要：

Objective：To investigate the physiological and pathological function of HSP27 by construct and identify the recombinant adenoviruses containing green fluorescence protein（GFP）and mouse HSP27 gene with PAdEasy system. Methods：The coding region of HSP27 was subcloned into the shuttle vector pAdTrack-CMV to form pAdTrack-CMV-HSP27. The identification was performed by PCR,sequencing and restriction digest. Chemical transformation of the plasmid pAdEasy-1 into E. coli BJ5183 strain was performed to prepare BJ5183-pAdEasy-1 as the competent bacterium,in which pAdEasy-1 and pAdTrack-CMV-HSP27 were cotransformed. Finally,the recombinant adenovirus containing the coding region of HSP27 gene was constructed by transfecting 293 cells with linearized adenoviral genomes of Ad-CMV-HSP27,and produce was used to infect mice spermatocyte,the expression of HSP＇27 protein was detected by Western blot. Results：The recombinant plasmids identified by PCR and directly sequencing were positive. GFP was observed after construction,high concentration of spermatocytes infected by recombinant adenortrus was harvested,and HSP27 protein expressed correctly. Conclusion：The recombinant adenoviruses expressing HSP27 and GFP using AdEasy system, which will facilitate further functional studies of HSP27, are successfully performed in this study.