Cloning and prokaryotic expression of actin related protein 2/3 complex subunit 4 of Taenia saginata asiatica

ObjectiveTo clone,express and conduct aprelimina ry mimuno reacticity study on genes of Taenia saginata asiaticas' act in related prote in 2/3 complex subunit 4(Arp2/3).MethodsBased on the Arp2/3 sequence template in the library of cDNA of Taenia asiatica,a pair of special primer was synthesized to amplify the gene through PCR technology.The genes were cloned in to prokaryotic expression vector Pet-28α(+)inducing the target genes to conduct expression through IPTG in competent Escherichia coli BL21/DE3 processed by CaCl₂.The products of the expression was identified with SDS-PAGE.As the expression took place in an inclusion body,SKL was used to achieve purified recombinant prote in and the immuno reacticity study was conducted with We stern blotting.ResultsPCR,double enzyme digestion and DNA sequencing indicated that pET 28α(+)and Arp2/3 recombinant plasmid was succe ssfully constructed.SDS-PAGE results showed that the gene expression took place in Escherichia coli BL21/DE3,and highly pure prote in was achieved after the dissolving,refolding and particle exchange chroma tography of inclusion body deposits.The recombinant prote inreacted with Taenia asiatica and taeniarhynchus saginatus infected patients' serum,which indicated the mimuno reacticity of the prote in.ConclusionThe Arp2/3 subun it 4 gene is successfully cloned.The recombinant prote in is obtained through expression and purification,and the genes' mimuno reacticity is confirmed,which provides the foundation for further studies of the gene.