Improved Recombinase Technology for Targeted Marker Free Integration and Founder Line Production for Risk Assessment
Project Number: 5325-21000-020-02
Start Date: Jun 01, 2011
End Date: Sep 30, 2013
The research objectives are to reduce potentially negative effects of transgene insertion and genomic presence. The aim of this proposed research is to investigate the use of novel unidirectional recombinases Bxb1, CinH, ParA and phiC31 to implement a Recombinase-Mediated Cassette Exchange (RMCE) technique for precise integration with simultaneous marker removal. The specific goals are: 1) To identify the most efficient pair of recombinase enzymes for dual unidirectional RMCE. 2) To demonstrate proof of concept with dual unidirectional RMCE in Glycine max. 3) To generate transgenic founder Glycine max lines containing the RMCE genetic platform for precise biotech risk assessment.
Single copy transgenic Glycine max founder lines will be generated containing the selection gene cassette flanked by fused recognition sites. An exchange vector will be biolistcally transformed into the various Glycine max founder lines. Recombinase mediated cassette exchange will be examined by negative selection and used to score the most efficient pairs. The most effective Glycine max founder lines and recombinase pairs will be published and made publicly available. Documents Trust with the United Soybean Board. Log 42344.