1a.Objectives (from AD-416):
Develop Potato Virus Y (PVY) resistance in existing cultivars by: 1)testing the S. hirsutum pot-1 gene as a source of PVY resistance in potato;. 2)doing a molecular characterization of eIF4E-transgenic lines; and. 3)produce minitubers for proof of concept field studies.
1b.Approach (from AD-416):
Inoculation studies will be done using Potato Virus Y O strain (PVYO) Potato virus Y NTN strain (PVYNTN), and Potato virus Y N:O strain (PVYN:O) strains in a greenhouse to assay the resistance in four cultivars transformed with the pot-1 gene. Molecular characterization of the transgenic lines will be assayed for sequence insertion and copy number using a Southern Blot analysis. Minitubers of the transgenic lines will be produced in sufficient numbers to allow replicated field trials for PVY resistance in 2012.
Fifteen transformed potato lines representing two standard cultivars (Russet Norkotah and Silverton Russet) were tested for PVY resistance by inoculating individual sets each with PVYO, PVYNTN, and PVYN:O strains. One of the eight Silverton lines showed resistance against all three virus strains tested. To-date, this experiment has confirmed that the transformations have produced one resistant line each of Russet Norkotah, Classic Russet, Silverton Russet, and one breeding line (MSE149-5Y). Currently, a trial using breeding lines is planted and is being tested under field conditions. Results from the field will be compared with the previous greenhouse tests. The inoculations in the field are being done with locally occurring field collected virus isolates. Michigan State is now isolating RNA from the transformed lines to look at the expression level of the eIF4E gene. Our hypothesis is that the higher gene expression level of eIF4E provides greater resistance to the three strains of PVY. We are also propagating the most resistant lines for greenhouse mini-tuber production so that field trials will be conducted in 2013. In those trials we will compare the transformed lines to the non-GM controls for both virus resistance and agronomic performance. We are continuing to conduct transformation experiments with Russet Norkotah line selections, but the efficiency of the regeneration for those lines is much lower. The research conducted contributes to Objectives 2 and 4 of the in-house project.