Sexually-mature mungbean (Vigna radiata (L.) Wilczek) plants were efficiently regenerated from cotyledonary node explants. The explants were capable of directly developing multiple shoots on basal media devoid of any growth regulators. The shoot multiplication was influenced by media composition, growth regulators, age of donor seedling and explant type. The explants with both the cotyledons attached to the embryonic axis excised from 4-d-old seedlings, produced the highest number of shoots (5 or 6) in 100% of the cultures within 2 weeks on B5 basal medium (BBM) containing BAP or 2-iP, respectively, (at 5x10(-7)M) and 3% sucrose. Shoots elongated and developed better using BAP. Increasing micronutrients, carbohydrate and nitrogen levels in the medium above the original formulation of B5 basal medium appeared to be of no benefit for increasing the number of shoots. The shoots were rooted on basal MS medium or MS containing 10(-6) of NAA, IAA or IBA. This protocol was found applicable to six other cultivars of mungbean. One hundred rooted shoots were successfully established in soil in the glasshouse, where 90% of them survived. The regenerated plants flowered precociously, but produced normal pods and viable seeds.
Conditions for plant regeneration from excised cotyledons of Vigna radiata were studied. Complete plant developed from the uncallused proximal ends of cotyledons on Murashige & Skoog's (MS), Gamborg's (Bs) and 'C' (MS salts + Bsvitamins) basal media. The basal medium 'C' was found to be best for plant regeneration. Regeneration frequency, however, varied with genotype, size, orientation and age of explant and the different plant growth regulators combination in the medium. Addition of cytokinins induced callusing at the proximal ends of cotyledons followed by multiple shoot formation. Out of 6-benzyl aminopurine (BAP), kinetin (KIN), N (A 2 isopentyl) adenine (2iP) and adenine sulphate (AS), only BAP and KIN were found to be more effective in enhancing the frequency of shoot regeneration. BAP at 1 x 10-SM induced maximum (60%) shoot regeneration whereas maximum number of shoots (8 to 9 shoots) per explant was observed with 5 x 10 6 M BAP. Cotyledons excised from two-day old seedlings were most regenerative. The regenerative response of cotyledons decreased when sliced into two equal parts either longitudinally or transversely. Callusing and organogenic differentiation occured only if the petiolar end of cotyledons was in contact with medium. None of the tested treatments were effective in inducing shoot bud differentiation from subcultured callus. Well developed shoots rooted when incubated on half strength MS, MS and MS basal medium supplemented with IAA (5 x 10-6M). The rooted plants were transferred to pots and later established in the field with 60% success.
A protocol for regeneration and micrografting of shoots of lentil (Lens culinaris Medik) was developed. Multiple shoots (4-5) were regenerated from cotyledonary node explants on Murashige and Skoog (MS) medium containing 8.8 mM 6-benzylaminopurine. In vitro regenerated shoots were micrografted on rootstocks with 96% efficiency. The successful grafts were transplanted to pots in Redi-earth TM , hardened off and were grown to maturity with 100% success. The success of the micrografting was independent of the nature and concentration of growth regulator used in shoot initiation medium and the time period for induction of shoots. The protocol was successful with several cultivars of lentil. The advantages of micrografting over in vitro rooting are discussed.
The effect of NaC1 stress on the activities of nitrate reductase (NR), glutamate dehydrogenase (GDH) and glutamate synthase (GOGAT) in callus lines of Vigna radiata which differ in salt resistance, was studied at weekly intervals upto 28 d of growth. After 28 d, the NaC1 resistant callus (selected at 300 mM NaC1) at NaCI concentrations higher than 200 mM maintained higher NR activity than non-selected line. NaC1 stress also affects aminating and deaminating activities of GDH. The NADH-GDH activity in the presence of NaC1 was higher in the resistant than nonselected line. On the other hand, NAD-GDH activity in both the lines was completely inhibited after 7 d of growth. The increased activity of NADH-GDH in resistant calli may play a vital role in protecting the cells from toxic effect of increased endogenous level of ammonia which probably accumulates due to efficient NO 3-reduction. NADH-GOGAT activity was found to decrease under salt stress in both the callus lines. Nitrogen assimilation in salt-resistant calli under salt stress was found to be characterized by high NR and NADH-GDH activities, concomitantly with low GOGAT activity.
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