A one-time inducible transposon for creating knockout mutants

Abstract

The maize transposon Ac can move to a new location within the genome to create knockout mutants in transgenic plants. In rice, Ac transposon is very active but sometimes undergoes further transposition and leaves an empty mutated gene. Therefore, we developed a one-time transposon system by locating one end of the transposon in the intron of the Ac transposase gene, which is under the control of the inducible promoter (PR-1a). Treatment with salicylic acid induced transposition of this transposon, COYA, leading to transposase gene breakage in exons. The progeny plants inheriting the transposition events become stable knockout mutants, because no functional transposase could be yielded. The behavior of COYA was analyzed in single-copy transgenic rice plants. We determined the expression of the modified transposase gene and its ability to trigger transposition events in transgenic rice plants. The COYA element thus exhibits potential for development of an inducible transposon system suitable for gene isolation in heterologous plant species.

The use of hygromycin phosphotransferase gene (hpt) with an artificial intron to obtain marker-off transgenic plants

“…Our results show that KH harbors similar transposition efficiency as previous PR-1a: TPase-based inducible transposon in rice (Charng et al, 2007). Taken our previous and current works together, we located the end of the inducible transposon in an intron of a target gene for subsequently removing its function in transgenic plants (Charng et al, 2008;Li et al, 2008;Tai et al, 2011). All these works are based on the fact that before transposition, the insertion of the end of transposon in an intron did not obviously affect the normal splicing process of the target gene.…”