Assessment of Bt cotton genotypes for the Cry1Ac transgene and its expression

Abstract

SUMMARYGenetically modified (GM) plants expressing Bt toxin provide protection against lepidopteran pests. The only GM crop in Pakistan is Bt cotton, which was illegally imported and adopted rapidly by cotton producers. Farmers gained access to the seed of many unapproved Bt genotypes before the matter was picked up and formal approval granted by the relevant governmental agencies. The present study was conducted to evaluate the samples of Bt cotton, collected from farmers and seed dealer, for transgene integration and expression. Seeds of 52 cotton genotypes, labelled as Bt, were collected from various farmers and seed dealers. An immunoblot strip test was carried out, which showed that only 0·86 of the samples collected were synthesizing Cry1Ac toxin. According to multiplexed polymerase chain reaction (PCR) results, 0·86 of the genotypes tested were positive for the Mon531 event (an ‘event’ is a specific genetic modification in a specific species) and 0·14 were negative for any transgene. Transcript analysis of transgenes in positive genotypes by real-time Rt-PCR confirmed the synthesis of mRNA in all genotypes but with significant variation. The concentration of Bt toxin revealed by enzyme linked immunosorbent assay (ELISA) showed that only 0·02 genotypes had the reported optimum level. The real-time PCR and ELISA results further confirmed the attenuation of transgene expression at transcriptional and translational level by various internal and external factors. The same type of event was found in all genotypes, with significant variation in toxin level, revealing the impact of genetic background on transgene expression. The findings support the recommendation to improve the existing quality criteria for transgenic cotton variety approval and certification in Pakistan, with the inclusion of toxin concentration in the list of parameters to be considered.