Regeneration of pea (Pisum sativum L. cv. Century) plants by in vitro culture of immature leaflets

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Abstract
In vitro regeneration of plants from immature leaflets of 3 day-old pea (Pisum sativum L. cv. Century) seedlings was studied under defined nutritional, hormonal and environmental conditions. Immature leaflets isolated from the second and third apical leaves of aseptically germinated seeds were cultured on MS medium containing vitamins as in B5 medium, 3% sucrose, 0.8% agar and supplemented with 0.1, 1, and 10 μM concentrations of naphthaleneacetic acid (NAA) and 1 and 10 μM levels of benzyladenine (BA) in various combinations. Shoot regeneration from the primary callus occurred within 45 to 90 days of culture in most of the hormone combinations. Although the number of calli producing shoots was maximal at 10 μM levels of NAA and BA, multiple shoot regeneration was predominant at a combination of 0.1 μM NAA and 10 μM BA. Indoleacetic acid (IAA) and kinetin (K), both at 10 μM, also induced shoot regeneration. No shoots were regenerated when 10 day-old leaflets were used as explants. Root production generally occurred on non-shoot regenerating calli. Roots were induced to differentiate by transferring the regenerated shoots onto half-strength B5 medium supplemented with 1 μM NAA.