PhD

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.