Rapid method for plasmid DNA recombination (Murakami-system)

Abstract

DNA recombination is a useful technology for cloning and subsequent functional analysis, while standard techniques for plasmid DNA recombination have remained unchanged. In the present study, we introduced rapid method for plasmid DNA recombination, which we named “Murakami-system”, to complete the experiments in under 33 h. For this purpose, we selected the following: PCR amplification with 25 cycles and E. coli strain with rapid growth (incubation time of 6–8 h). In addition, we selected rapid plasmid DNA purification (mini-prep; ∼10 min) and rapid restriction enzyme incubation (20 min). This recombination system enabled rapid plasmid DNA recombination within 24–33 h, which could be useful in various fields. We also established a 1-day method for competent cell preparation. Our rapid recombination system allowed several sessions of plasmid DNA recombination to be performed every week, which improves the functional analysis of various genes. • “Rapid method for plasmid DNA recombination (Murakami-system). • E. coli strain with rapid growth (incubation time of 6–8 h). • Combination of rapid protocols (PCR, electrophoresis, DNA purification, ligation, and mini-prep) enabled plasmid DNA recombination within 24–33 h.

Keywords

• Rapid method for plasmid DNA recombination (Murakami-system)