Development of an Efficient Method to Enhance Regeneration Capacity Using Different Culture Conditions and Explants in Bread Wheat (Triticumaestivum L.)

Abstract

In vitro mutagenesis is a novel tool for improvement and development of new plant genotypes. The purpose of present investigation was to develop an efficient method of enhancing the regeneration capacity in wheat. The effect of plant growth hormones (PGR) has been assessed on callus induction response and regeneration frequency. The highest callus induction with lowest precocious germination recorded in genotypes viz. C306 and HD-2967 genotype and highest shoot regeneration and better number of roots among both genotypes. The healthy wheat plantlets were transferred to the plastic pots for acclimatization. The mature embryo of wheat seeds treated with EMS and calluses treated with UV light of two genotypes C-306 and HD-2967. The mutant lines of genotype C-306 showed highest callus induction with low precocious germination in treatment T-6 (0.5% EMS for 2 hours). The highest callus induction was found mutant genotype HD-2967 with low precocious germination with treatment T-5 (0.5% EMS for 1 hour). In highest shoot regeneration observed in treatment T-4 (0.5% EMS for 30 minute) with 1.5 shoot per callus in mutant plantlet of genotype C306 and highest shoot regeneration observed treatment T-5 (0.5% EMS for 1 hours) with 1.4 shoot per callus. In case of UV light treated mutant lines of genotype C-306 showed 73.8 percent shoot regeneration with 0.7 shoot per explants when treated with UV-3 (UV light for 2 hours) and lines of genotype HD-2967 showed 82.2 percent shoot regeneration with 8.6 shoot per explants when treated with UV-2 (UV light for 1 hours) were observed statistically significant. Molecular analysis was done to see whether any change occurred by treatment of mutagens (EMS and UV light) in regenerated plants of wheat. For this purpose SSR and ISSR primers were used to analyze the regenerated normal and mutant plants of the twenty six normal and mutant plants. EMS concentration (0.5% EMS for 2 hour) did mutation and mutate on 5A, 7A, 7B and 4D chromosomes of regenerated lines at site of SSR marker loci. In case of UV light concentration (UV light for 30 minute) mutation occurred on 1B, 5A and 7B chromosomes in wheat regenerated lines at site of SSR marker loci in both the genotypes Viz. C306 and HD2967. The mostly mutant plantlet observed EMS concentration (0.5%EMS for 2 hour) causes mutation in genotype C306 on different ISSR marker loci. In case of UV light concentration (UV light for 30 minute), it causes mutation in both genotypes Viz. C306 and HD2967 at a site of ISSR marker loci. In SSR analysis PIC values varied from 0.00 (Xgwm 190-5D) to 0.946 (Xgwm 60-7A) with an average of 0.620. In ISSR analysis PIC values varied from 0.103 (UBC 826) to 0.946 (UBC 876) with an average of 0.744. The present study would be useful in developing transgenic lines for improved traits in future.