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Subject: Vegetable Science × Author: Manpreet Kaur × End date: 2019 × Start date: 2012 × Thesis Type: M.Sc ×
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    ThesisItemOpen Access
    Standardization of thermotherapy and tissue culture techniques for virus free garlic (Allium sativum L.)
    (Punjab Agricultural University, Ludhiana, 2018) Manpreet Kaur; Sharma, Madhu
    Garlic (Allium sativum L.), being an agamic crop lacks seed production and is mainly propagated vegetatively. This phenomenon of vegetatively propagation leads to degeneration of varieties over the generations in terms of production and productivity through buildup of virus complexes. Consequently, to develop vigorous and virus free plants, the present investigation entitled “Standardization of thermotherapy and tissue culture techniques for virus free garlic (Allium sativum L.)”. In this regard thermotherapy, tissue culture and combination of both in vivo and in vitro techniques were exploited and standardized to obtain virus free plants in garlic. 200 virus infected bulbs of G-282 based on morphological symptoms were harvested and subjected to DAS-ELISA for the confirmation of potyvirus. Cloves were exposed to hot air and hot water ranging from 40-70oC and 55-85oC, respectively, for estimating per cent survival rate and days taken for germination. Treatments exhibiting 80-100% survival rate were used for development of virus free plants. Hot air at 55oC for 20 hours and 60oC for 30 minutes showed 80-100% survival rate in 13-21 days with 100% virus elimination whereas, 60oC for 15 minutes resulted in 75% virus free plants. Hot water at 55oC for 10 minutes showed 90% survival rate with 66.67% virus elimination whereas 55oC for 20 minutes, and 65oC for 10, 20 & 30 minutes gave 100% virus free plants. In in vitro regeneration, shoot tips and stem discs of different genotypes viz., G-282, AC-102 and PG-20 were excised from surface-sterilized virus infected cloves and cultured on MS medium supplemented with growth regulators for shoot regeneration. In this study, shoot tip and stem disc culture gave 85% and 75% shoot regeneration under in vitro conditions, respectively. In shoot tip culture, more average number of shoots (3.1) with greater average shoot length (9.49 cm) was found on MS medium supplemented with NAA and BAP 0.2 mg/l each whereas, stem disc culture showed less average number of shoots (3) with 9.19 cm average shoot length. G-282 and AC-102 were found more responsive for in vitro regeneration through shoot tip. However, AC-102 and PG-20 were found recalcitrant for stem disc culture, thereby exhibiting genotypic differences in regeneration among the cultivars. Stem disc showed more regeneration in MS medium supplemented with 1.5 mg/l of BAP as compared to 1 mg/l BAP. Higher percentage of root induction in shoot tip culture was initiated from ½MS medium with 30 g/l sucrose. 80% and 25% regenerated plants were found to free of potyvirus infection in shoot tip and stem disc culture, respectively as indexed by RT-PCR. Suggesting shoot tip to be a better approach than stem disc in garlic. In combination of in vivo and in vitro techniques, cloves were subjected to hot air and hot water prior to excising shoot tips. Pre-culture exposure to hot air at 40oC for 10 & 20 days and 55oC for 20 hours showed 40-60% regeneration rate with 100% virus elimination whereas 60oC for 15 minutes regenerated 66.67% virus free plants. In hot water, 55oC for 10 and 20 minutes gave 80 and 50% regeneration with 100% virus elimination. Cloves exposed to hot water at 65oC did not responded in in vitro conditions. The regeneration of virus free plants through tissue culture techniques requires technical skills and is a costly venture. Thus, exposure of garlic cloves to hot air at 60oC for 30 minutes or hot water at 65oC for 20 minutes found efficient and cost effective approaches to produce virus free plants.