In vitro mutagenesis of Chrysanthemum morifolium cultivars using ethylmethanesulphonate (EMS) and mutation assessment by ISSR and IRAP markers

Mutation induction is a feasible and established breeding method for crop improvement and genetic diversity creation to introduce new plant cultivars. The present study was aimed at mutagenesis of four chrysanthemum cultivars (‘Homa’, ‘Fariba2’, ‘Arina’, and ‘Delkash’) using ethyl methanesulfonate (EMS) (0, 0.125, 0.25, and 0.5%) as mutagen and leaf disks as explants to obtain novel variants. In addition, genetic polymorphism among mutants and their parents was detected using inter simple sequence repeat (ISSR) and inter-retrotransposon amplified polymorphism (IRAP) molecular markers. A total of 2082 plantlets were produced through EMS induced mutagenesis under in vitro conditions and at the end 58 mutants including 28 leaf and 32 flower mutants were analyzed for phenotypic and molecular variation. The explant survival rate decreased by increasing EMS concentration. A wide range of phenotypic leaf and inflorescence variability was obtained in four studied chrysanthemum cultivars confirming the efficiency of EMS to create genetic variation and desired mutants. All generated variants with different inflorescence and leaf shape and color were maintained through cuttings and they expressed same traits in the next generation. The mutants were different in leaf size and shape, plant height, day to flowering, inflorescence head size, ray floret color and ray floret size. The used ISSR and IRAP primers could classify chrysanthemum mutants based on cultivar and somewhat based on used EMS concentration confirming their effectiveness for the discrimination of real variants that allow their earlier selection and reduction of the mutant population size. The in vitro EMS-induced mutation can be a promising tool to assist breeding programs for the generation of new chrysanthemum cultivars.