Assessment and induction of variability through in vitro mutagenesis in cassava (Manihot esculenta Crantz)

Abstract

The study entitled “Assessment and induction of variability through in vitro mutagenesis in cassava (Manihot esculenta, Crantz.)” was carried out between 2012 and 2014 in the Department of Plant Breeding and Genetics, College of Horticulture, Vellanikkara. The objective of the study was to assess the genetic variability in the short duration cassava germplasm and induction of variability through in vitro mutagenesis in selected genotypes. Field evaluation, standardization of protocols for in vitro regeneration, in vitro mutagenesis and assessment of variability of in vitro mutated plants were done. Wide genetic variability existed among the collected short duration cassava genotypes. The colour of petiole and root cortex was found to be the most variable qualitative trait for above ground and tuber portions respectively. High magnitude of phenotypic and genotypic coefficient of variation along with high heritability and high genetic gain was observed for branch number, branch height, tuber fresh weight, and Cassava Mosaic Disease. All traits except tuber neck, branch number, scar number and internode length were positively correlated with fresh tuber yield. High direct contribution towards tuber yield was exerted by shoot biomass, tuber dry matter content and harvest index indicating that these are reliable predictor variables for increased yield. Among the biochemical traits, high heritability and high genetic gain was observed for HCN content, amylose content and starch content. Biochemical analysis indicated the occurrence of high starch genotypes (Sree Jaya, CC10 and CC7) suitable for industrial starch production. Vellayani Hraswa with lower starch content was more suited for food or feed purposes, than industrial purposes. The genotype CC10 with easiness in peeling, good taste after cooking and less cooking time scored maximum in organoleptic evaluation. Sree Jaya was sweet on chewing, had highest starch content, lowest HCN content and less fibre content. Diversity analysis indicated that the cassava genotypes grouped into five clusters. No parallelism was found to occur between geographic distribution and genetic diversity. Selection index constructed for the identification of the best genotypes indicated that CC1 and Sree Jaya were the most promising genotypes. CC1, a farmers’ variety from Malappuram district was found to be the best genotype with respect to yield and resistance to CMD, but with a comparatively high HCN content. In vitro mutagenesis in cassava was done using the genotypes CC1 and Sree Jaya. Sterilization of cassava nodal and leaf explants was accomplished by washing with 5 per cent Teepol solution for two minutes, followed by washing for one minute with 75 per cent ethanol and washing for one minute with 0.05 per cent solution of mercuric chloride. Friable embryogenic callus (FEC) for both CC1 and Sree Jaya genotypes was obtained from immature leaf explants cultured in MS media with 3.0 per cent sucrose (MS3), either with 6.0 to 8.0 mg l-1 of 2,4- D , or with 1.0 mg l-1 BAP + 0.2 to 0.5 mg l-1 NAA. Somatic embryos for both genotypes were obtained from FEC cultured in media MS3 with 8.0 to 10.0 mg l-1 picloram and germinated into the plantlets on MS3 media with 4.0 mg l-1 BA or 0.25 mg l-1 TDZ. In vitro regeneration and multiplication from nodal explants were obtained in MS3 media containing either 0.25 mg l-1 TDZ or 2.0 mg l-1 BAP. Rooting of the in vitro plantlets was obtained in MS3 + 0.25 mg l-1 TDZ or 1/2MS1. The LD 50 value varied with the cultures used for in vitro mutagenesis. The LD 50 value for gamma radiation was 40 Gy, 30 Gy and 50 Gy for FEC, somatic embryoids and plantlets, respectively. LD 50 value for EMS was 1.20 per cent for FEC and somatic embryoids and 0.90 per cent for plantlets. Variation in response to mutagenesis was also observed between the two genotypes subjected to in vitro mutagenesis. There was significant difference in the growth characteristics of the mutagen treated in vitro cultures in both genotypes. Reduction of the number of shoots and leaves were more in CC1 compared to Sree Jaya. A combination of SoilriteTM with pure sand at 1:1 proportion was the best substrate for acclimatization of the plantlets outside the tissue culture lab. Fan and pad green house was the best structure for in vitro acclimatization of plantlets resulting in 47 per cent of success rate. Variability with respect to quantitative traits like height, number of shoots and number of leaves was observed in vitro plantlets in the hardening stage. The qualitative traits like colour of the petiole, stipule, emerging leaf and of the stem and the shape of central lobe of leaves varied between the mutated plants.