Genetic Analysis of Cotton Genepool revealed by RAPD - PCR

Abstract

The study of genetics and evaluation of Gossypium is important for improvement in the existing gene pool. Genetic variations within and among the species and cultivars of Gossypium showing different polymorphisms were studied using the Random Amplification Polymorphic DNA (RAPD) method. In order to achieve this it had first to optimize the methods by testing different conditions of DNA extraction, amplification and electrophoresis. 10-mer oligonucleotides were randomly chosen from commercial kits and employed as primers to amplify segments of the genome and the products were detected on agarose gels. DNA amplification parameters (Buffer composition, MgCl2, DNA template, primer, dNTPs, Taq DNA polymerase, sample volume, thermocycler program) were optimized to give reproducible bands. A total of 60 cotton genotypes were used to analyze the genetic diversity. Morphological characters of 13 species were studied and a dichotomous key was prepared on the bases of these characters. Twenty species of Gossypium belonging to different genomes (A-F) were analyzed by using 63 primers. The similarity coefficients ranged from 0.03 to 0.64. Maximum similarity 64% was observed between G.herbaceum and G.arboreum and lowest similarity 3% was observed between G.stocksii and G.longicalyx. The dendrogram based on similarity coefficients was constructed by using the Unweighted Pair Group Method with Arithmetic Averages (UPGMA). Twenty species of Gossypium clustered into five distinct groups, and the two species G. longicalyx and G. stocksii stands separately in the dendrogram. Twenty cultivars of G. hirsutum were analyzed by using 75 primers. The similarity coefficients ranged from 0.53 to 0.92. Maximum similarity 92% was observed between CIM-1100 and CIM-446 and the exotic cultivars showed 53% genetic similarity with the local cultivars of G. hirsutum. The dendrogram verified that the bulk of the genotypes were clustered in five groups. To evaluate the stability of cultivars, variation within cultivars (CIM-473, CIM-448, CIM-1100, NIAB-78) was studied. Maximum similarity within the cultivars, 92- 100% was observed in NIAB-78 and minimum similarity, 80-92% was observed in CIM-1100 by using 40, 10-mer primers. Similar amplification profiles were observed in four different cultivars. Cultivars of G. hirsutum were more closely related to each other than to the wild species of Gossypium.