Transcription–replication collision increases recombination efficiency between plasmids

• Recombination induced by transcription–replication collision (TRC) was studied.
• A TRC repair plasmid was co-transfected with a mutant homologous plasmid.
• TRC improved the recombination rate between the plasmids.
• The improved recombination rate was mainly due to homologous recombination.
• The recovered plasmids were transformed into Escherichia coli, which were plated on an ampicillin-resistant plate.

It has been proposed that the stalling of the replication forks can induce homologous recombination in several organisms, and that arrested replication forks may offer nuclease targets, thereby providing a substrate for proteins involved in double-strand repair. In this article, we constructed a plasmid with the potential for transcription–replication collision (TRC), in which DNA replication and RNA transcription occur on the same DNA template simultaneously. Theoretically, transcription will impede DNA replication and increase homologous recombination. To validate this hypothesis, another plasmid was constructed that contained a homologous sequence with the exception of some mutated sites. Co-transfection of these two plasmids into 293T cells resulted in increased recombination frequency. The ratio of these two plasmids also affected the recombination frequency. Moreover, we found high expression levels of RAD51, which indicated that the increase in the recombination rate was probably via the homologous recombination pathway. These results indicate that mutant genes in plasmids can be repaired by TRC-induced recombination.