Development of a polymerase chain reaction assay for detection of three canola transgenes

Many countries are developing or implementing regulatory requirements to monitor for the presence of genetically modified (GM) materials in seeds, grain, and derived food products using DNA and protein-based methods. There is no published report on the detection of different GM transgenes in canola, and this study is aimed at developing qualitative PCR methods for the three major GM transgenes commercially available in canola. Primer sequences were generated from GenBank and previously published information to develop a polymerase chain reaction (PCR) detection method for Roundup Ready (glyphosate tolerance, GT73 event), Liberty Link (glufosinate ammonium tolerance, HCN92 event), and BX (Bromoxynil tolerance, OXY235 event) canola varieties. On using PCR, two primer pairs for each of the GT73 and HCN92 and one primer pair for OXY235 amplified specific amplicons for the three GM transgenes. All three GM transgenes were detected simultaneously by multiplexing the five primer pairs in a single PCR reaction. Multiplexing of the five primer pairs for DNA prepared at 1% (one GM seed in 99 non-GM seeds) and 0.5% (one GM seed in 199 non-GM seeds) levels generated the expected DNA fragments for GT73, HCN92, and OXY235. This information will lay the groundwork for the development of a quantitative PCR assay for canola transgenic events.