Expression of GFP and Bt transgenes in Brassica napus and hybridization with Brassica rapa

Abstract

It is possible to monitor the movement of transgenes by tagging them with green fluorescent pro- tein (GFP). In order to develop a model to study trans- gene flow, canola (Brassica napus cv Westar) was trans- formed with two GFP constructs, mGFP5er (GFP only) and pSAM 12 (GFP linked to a synthetic Bacillus thuringiensis (Bt) cry1Ac endotoxin gene). Transformed callus sectors that fluoresced green were preferentially selected in the tissue culture process. Four independent GFP canola events and 12 events of GFP/Bt canola were regenerated through tissue culture. GFP fluorescence was macroscopically detectable throughout the entire life cycle of canola. The GFP/Bt events were insecticidal to neonate corn earworm (Helicoverpa zea) larvae and prevented herbivory damage. Fluorescence intensity at 508 nm varied between the independent transformation events, and ranged from 7.6× 10 5 to 13.8× 10 5 (counts per second) in contrast with the wild-type at 5.3 × 105 cps. Nine GFP/Bt and three GFP events were hybridized with three wild accessions of B. rapa. The resultant hybrids fluoresced green and were insecticidal to neonate corn earworm larvae to the same degree as the transgenic ca- nola parents. However, fluorescence intensities of the hemizygous F 1 hybrid lines were lower than the respec- tive original homozygous canola parents. Each F 1 hybrid line was backcrossed by hand onto the B. rapa parent, and transgenic backcrosses were produced at rates rang- ing from 15% to 34%. These data suggest that GFP can be used as a tool to monitor transgene flow from crop species to wild relatives.