Identification of immune system related mutant genes in cotton leaf curl virus disease (CLCuD) resistant and susceptible M5 cotton (Gossypium hirsutum L.) plants
2016
Majeed, M.
Cotton is the most important fiber as well as an oilseed crop of Pakistan. Cotton leaf curl virus disease (CLCuD) is a big threat to the cotton production in the country. Classical breeding approaches for the production of CLCuD resistant varieties are not satisfactory because of long generation period, emergence of new Cotton leaf curl virus (CLCuV) strains and large genome size of cotton (Gossypium hirsutum L.). In this situation, mutation is a useful source to create genetic variability for resistance against this fatal disease. Spontaneous mutation occurs at very low rate so induce mutation can be used to generate useful genetic variations and crop improvement. Therefore, in this study an experiment was conducted on earlier screened CLCuD resistant and susceptible plants of M5 cotton population for mutation identification. Fresh leaves of twenty resistant and susceptible genotypes were collected for DNA extraction and identification of mutated defense related gene families (DGAs and RGAs). The DNA, isolated from mutant and wild plants (PB-899) were mixed in equal concentration and subjected to PCR amplification. Heteroduplexes were formed and cleaved by CEL 1 nuclease. CEL 1 treated samples were resolved on 2% agarose gel to detect the mutations. Different bands of hetroduplexes were identified in meloidogyne-induced cotton-3 gene (MIC-3 gene) of primer DGA 5 and NAC 5 gene, which were amplified with two primer pairs DGA 3 and DGA 4. The identified mutation of the resistant gene family in those plants, who have already been screened out as resistant, can open new horizon in defining the role of these genes for resistance development.
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