Lentiviral gene vector transformation in Escherichia coliderived DH۵α, JM۱۰۹ and Stbl۴ cells

Background and Aim : Bacterial cell transformation is essential for gene cloning. Althoughplasmid gene transfer recombinant lentivectors are highly effective, they do not transform readilyin most cells. E coli derived Stbl۲ and Stbl۳ strains maintain structure of lentiviral plasmidpRRL.SIN.cPPT.PGK/Oligo۲-IRES-DsRedWPRE and boost colony growth. However, Stbl۴appears to be even more efficient to justify further study. We thus investigated plasmidtransformation in DH۵α, JM۱۰۹ and Stbl۴ cells derived from Escherichia coli.Methods : Transformation efficiency was calculated based on the method of Tu et al. (۱۴). by thefollowing two equations: Number of transformed colonies (cfu) = number of bacterial colonies ×dilution index× volume of transformed mixture ÷ volume taken to the plate. Transformingefficiency = number of transformed colonies ÷ plasmid concentration in micrograms.Results : After electrophoresis of recombinant plasmids in different dilutions, plasmidconcentration was determined and concentration was adjusted to ۱۰۰۰ ng / ml. The results ofplasmid electrophoresis can be seen in Figure ۱.Conclusion : Overall, the efficiency of the transformation in the present study for DH۵α, JM۱۰۹and Stbl۴ averaged ۰.۲۶ * ۱۰۶ , ۰.۸۹ * ۱۰۶ and ۱.۵۷ * ۱۰۶ cfu /μg, respectively which indicatedthat Stbl۴ was a more suitable competent cell for recombinant pCDH vectors.

کلیدواژه ها:

Transformation ، competent cell ، DH۵α ، JM۱۰۹ ، Stbl۴ ،