Precision Mutagenesis in Plants via Genome Editing

There are two types of genome editing technology, targeted mutagenesis and gene targeting. Targeted mutagenesis is useful if we want to knock-out a targeted gene. In contrast, gene targeting enables precise modification of the targeted gene using a specific template DNA. In both cases, we would like to remove transgne used for genome editing completely from the plant genome to leave a desired point mutation.

In the case of a targeted mutagenesis in seed-propagated crops, once integrated into the genome as a transgene, sequence-specific nucleases (SSNs) genes can be segregated out by self-pollination or back-crossing. However, in the case of crops that are propagated vegetatively, we would like to remove transgene without using genetic segregation. We found that piggyBack transposon can be used as a reversible transgenesis vector in plants to put SSNs IN and OUT without leaving any footprints.

Usually, a positive selection marker gene is left at the targeted gene after positive/negative-selection-mediated gene targeting. Recently, we succeeded in removing the positive selection marker gene completely using a piggyBack transposition. We can modify any sequence in the rice genome precisely using this system.