Sequential recombination of SIV envelope protein (ENV) gene and study on transfection by the mutant clones
Objective To study the relationship between DNA fragments in ENV gene of SIVmac and viral replicating ability. Methods Reciprocal recombinants were constructed by replacing the clonable DNA fragments of ENV region of the nonpathogenic SIVmac142 clone with the pathogenic SIVmac 239 clone. Following confirmation by RFLP and partial sequence analysis, the aliquot mutant viruses (3μg) were used to transfect CEM×174 cell line, and the concentration of SIVmac main core protein(P27) in culture supernatants was closely monitored by enzyme-linked immunosorbent assay (ELISA) in order to identify the replicating ability of the mutants. Results Compared with SIVmac 239 clone, SIVmac 239 envGP recombinant (SIVmac142/239envGP/142) was still imbued with high replicating activity; and SIVmac239gp41 (SIVmac142/239gp41/142) or SIVmac239N-gp41 (SIVmac142/239N-gp41/142) with comparably decreasing activity, while SIVmac239C-gp41(SIVmac142/239C-gp41/142) lacked replicating activity. Conclusion ENV is an important regulating element to the replication or virulence of SIVmac239 clone.