Development of Doubled Haploid Lines for Blast Resistance in Rice Oryza Sativa L through Anther Culture

Abstract

Rice (Oryza sativa L.) is the second most important food crop of the world. India is the second largest rice growing country after China. Among biotic stresses, blast caused by Magnaporthe oryzae is one of the most serious production constraints in almost all the rice growing ecologies of the world. Production of doubled haploids (DHs) through anther/microspore culture is a rapid approach to attain homozygosity that can shorten the time required to develop a cultivar. newline The present investigation entitled Development of Doubled Haploid Lines for Blast Resistance in Rice (Oryza sativa L.) through Anther Culture was carried out: to optimize media and culture conditions to enhance androgenesis in rice genotypes; development of doubled haploid lines through anther culture; and genotyping of doubled haploid population using SSR markers linked to blast resistance genes. To accomplish this, two lines (RML 22 and DHMAS), which had expressed resistant response against prevalent strains of Magnaporthe oryzae, were crossed as male parents with each blast susceptible lines viz. K448, K39 and K343 respectively for generating F1 population which were subsequently subjected to anther culture for development of doubled haploids. A healthy crop of F1 crosses along with female parents was raised to collect anthers at appropriate stage of pollen development. newline Callus induction potential of anthers in different genotypes could be enhanced many folds by supplementing amino acids, particularly a combination of cysteine and tryptophan, in N6-13 medium [N6 (3.97 g l-1) + Maltose (40 g l-1) + 2,4-D (2.5 mg l-1) + Kinetin (0.5 mg l-1) + tryptophan (25 mg l-1) + cysteine (40 mg l-1)]. High scores of callus and haploid regeneration frequencies were obtained in agar (0.8%) solidified MS-4 medium [MS (4.41 g l-1) + Sucrose (30 g l-1) + BAP (2.0 mg l-1) +Kinetin (0.5 mg l-1) + NAA (0.5 mg l-1)]. This medium could be refined further to achieve maximum regeneration of calli and haploids in amino acid supplemented MS-14 medium [MS (4.41 g l-1) +