In vitro shoot regeneration from cotyledonary leaf explant in chilli and bio-hardening of plantlets

A protocol for direct shoot regeneration from cotyledonary leaf explant in chilli was developed and method for bio-hardening of in vitro regenerated plantlets using Glomus mosseae, Gigaspora margarita and mixed arbuscular mycorrhizal fungi (AMF) strains were standardized. The experiment was undertaken with four chilli cultivars namely KtPL-19, Pusa Sadabahar, ArCH-001 and Salem. Explants were excised from 21-day-old in vitro raised seedlings. Direct shoot organogenesis was observed on cotyledonary explant with slight callusing. Number of shoot buds per explant was maximum (5.73) and days taken for shoot bud induction was minimum on MS medium supplemented with 1.0 mg l−1 of TDZ in all the cultivars. However, response of Pusa Sadabahar was better in terms of number of shoot buds than other cultivars under study. Fifteen to 20 days were required for shoot bud induction in all the cultivars except Salem which took 25–30 days. Among the different treatments tested for shoot multiplication the best treatment was MS + 6.0 mg l−1 BAP + 1.0 mg l−1 kinetin + 0.5 mg l−1 GA3. The length of the shoot increased with increase in BAP and GA3 levels. The in vitro regenerated shoots were inoculated on to half-strength MS medium supplemented with 1.0 mg l−1 IBA where more than 90 per cent rooting was observed. When in vitro raised plantlets were treated with AMF high plant survival was observed. The maximum survival (97.08%) was recorded with mixed strain of Glomus mosseae and Gigaspora margarita. The root and shoot length was also maximum when plantlets were treated with mixed AMF strains. The developed protocol may be used for mass multiplication of elite chilli genotypes as well as regeneration of genetically transformed cell/tissue.