MOLECULAR ANALYSIS AND DIFFERENTIATION OF GENETICALLY MODIFIED COTTON GENOTYPES

Abstract

The polymerase chain reaction (PCR) technique is a reliable and accurate tool to prove that plants may be genetically modified (GM). For ecological and regulatory reasons, it is important to know whether cotton (Gossypium hirsutum L.) varieties planted in Mexico are conventional or GM. Unequivocal procedures are necessary to identify them. Under the hypothesis that the presence of at least one transgenesis element would allow genotypic differentiation and clustering in blind samples; the objective of this study was to determine by PCR the presence of the transgenesis elements CaMV35S, nptII or Tnos in a set of blind cotton samples assumed to be GM, in order to differentiate clusters based on their genetic structure. The material studied was 20 cotton samples collected in the Comarca Lagunera, Mexico. Samples were analyzed with PCR to detect the type of regulatory transgenesis sequence used; then, the samples were genotyped with Inter-Single Sequence Repeat (ISSR) markers to determine type and number of clusters. The results indicated that 50% of the samples contained CaMV35S, nptII and Tnos sequences; 15% corresponding to nptII and Tnos; 5% to CaMV35S and nptII, and 5% to nptII only, and revealed that the gene construction in these cotton materials was distinct. Twenty-five percent of the samples tested had no transgenesis elements. The ISSR primers amplified 283 fragments, 61.5% of which were polymorphic. According to the values of the index of marker and polymorphic bands, the primers MicroAnch 4, MicroAnch 6 and UBC 872 were the most efficient in differentiating the samples under study. Cluster analysis and principal coordinate analysis separated the 20 samples into two clusters, in addition to the wild genotype; and analysis of molecular variance indicated that 85% of the variation was attributed to genotypes within the clusters. PCR is a useful molecular technique to identify transgenesis elements for grouping plants according to their genetic similarity.