Location: Crop Improvement and Genetics Research
Project Number: 2030-21220-003-002-T
Project Type: Trust Fund Cooperative Agreement
Start Date: Nov 1, 2019
End Date: Oct 31, 2024
Objective:
The goal of this project is to create soybean founder lines that are competent for genome targeted insertion via RMCE. USDA to establish a minimum of 3-5, single copy “founder lines” where the marker used for transformation can be excised by a recombinase and an additional site for stacking of a second cassette. Within these founder lines, there must be at least two genes – the selectable marker gene and another “gene of interest” (GOI-GUS) that can easily be assayed for activity.
Approach:
The lines will be analyzed for expression of the selectable marker gene and the GOI-LUC in the T0 generation and then will be followed in T1 for the hemizygous lines (minimum of 5 plants/line at the hemi stages); 15-25 analyzed per line. 2-3 of the founder lines will be “supertransformed-RMCE1” with a construct containing a second plant selectable marker and a second GOI. A minimum of 3-5 T0 “sibs” will be generated from each founder line where there is the presence of only the recombined second cassette. Analysis of T1 will be performed of the selectable marker gene and all GOI for these plants. 15-25 analyzed. 2-3 of the founder lines will be “supertransformed-RMCE2” with a construct containing the first plant selectable marker (same as original founder lines) and a third GOI-REN. A minimum of 3-5 T0 “sibs” will be generated from each founder line where there is the presence of only the recombined third cassette. Time permitting - analysis of the T1 will be performed of the selectable marker gene and all GOI for these plants. 15-25 analyzed.
