Construction and Identification of Recombinant Adenovirus Vector of siRNA Targeting Hsp27

Hsp27 is a member of the heat shock protein family,which plays important roles in a variety of physiological processes including protein chaperoning,steroidogenesis and especially protection against apoptosis.To construct an adenovirus vector that expresses small interfering RNA(siRNA) against Hsp27 to shut down its gene expression,the Hsp27 template DNA sequence was designed by using online tools.The sense and antisense siRNA oligonucleotide templates were chemically synthesized,annealed and cloned into adenoviral shuttle vector pShuttle-H1.The recombinant adenovirus vector was obtained by homologous recombination with pAdeasy-1 in bacteria BJ5183.The recombined adenovirus was produced in 293 cells and subsequently infected AD-293 cells by co-transfect with adenovirus vector expressing Hsp27.The adenovirus vector expressing small interfering RNA for Hsp27 gene could specifically shutdown the 67% Hsp27 protein by western blot.The recombinant adenoviruses expressing small interfering RNA forHsp27 gene was constructed successfully.The adenovirus vector can effectively down-regulate Hsp27 expression by co-transfect with adenovirus vector expressing Hsp27 in the present study,which will facilitate further functional studies of Hsp27.