Decrease in gyrase A protein expression in E. coli cells inhibited by antisense ribozymes

RNase P complexed with external guide sequence (EGS) represents a novel nucleic-acid-based gene interference approach to modulate gene expression. Nucleic acid-based gene interference technologies represent promising strategies for specific inhibition of mRNA sequences of choice. Recently, small interfering RNAs have been implicated in inducing endogenous RNase of the RNA-induced silencing complex in the RNA interference pathway to inhibit gene expression and growth of several human viruses. We report down regulation of protein expression of E. coli gyrase A, an essential gene for DNA supercoiling and antibiotic susceptibility in BL21 (DE3) strain of E. coli , using Ribonuclease P based external guide sequence (EGS) technique. EGS directed against gyrase A gene that was cloned into pUC vector, which contains the ampicillin (Amp) resistance gene. The recombinant plasmid pT7EGyrA was transformed into BL21 (DE3) and inductions were performed using IPTG. Western blot was done to investigate the downregulation of gyrase A protein. The results showed a significant decrease of gyrase A suggesting the utility of EGS RNAs in gene therapy applications, by inhibiting the expression of essential proteins.