Qualitative and Quantitative PCR-Based Detection Methods for Authorized Genetically Modified Cotton Events in India

Qualitative diagnostics for all five commercialized genetically modified (GM) cotton events for insect resistance in India is being reported for the first time in this paper. The cost-effective and robust multiplex PCR (MPCR)-based detection assay, distinguishing the insect resistant transgenic Bt cotton events, viz., MON531, MON15985, Event 1, GFM-cry1A, and MLS-9124, has been developed. This decaplex PCR assay targets nine transgenic elements, viz., sequences of four transgenes, three transgene constructs, and two event-specific sequences along with one endogenous reference gene. The LOD of the qualitative MPCR assay was up to 0.1%. A quantitative detection method for four widely commercially cultivated GM cotton events, namely, MON531, MON15985, Event 1, and GFM-cry1A, covering 99.5% of the total area under GM cultivation in the country, is also reported. A construct-specific real-time PCR assay has been developed for quantification of these GM cotton events with LOQ <0.05% and LOD <0.025%. The developed assays will be of great use to screen for the presence/absence of authorized GM cotton events in unknown samples and to check the authenticity of GM cotton seed samples.