title: A reproducible procedure for plant regeneration from seedling hypocotyl protoplasts of Vigna sublobata L.
abstract: Viable protoplasts of Vigna sublobata L. were isolated enzymatically from hypocotyls of axenic seedlings. Protoplast yields were dependent upon seedling age, with maximum yields (2.25 ± 0.35 × 10(6) g fwt(-1)) from seedlings aged 6 d. Protoplasts regenerated cell walls and underwent sustained divisions when cultured in either agarose-solidified or liquid K8P medium. The plating density affected the division frequency and plating efficiency; the division frequency (68 ±0 6.0%) was maximum at 4.0 × 10(4) ml(-1) while plating efficiency was maximum (1.3 ± 0.1%) at 5.0 × 10(4) ml(-1). Dividing protoplasts developed into microcalli, which produced glossy green compact nodular calli on transfer to 8.0 gl(-1) w/v agar-solidified medium containing MS salts, B5 organic components, 30 g l(-1) sucrose, NAA (0.2-0.5 mg l(-1)), zeatin riboside (0.5-2.0 mg l(-1)) and GA3 (0.5-1.0 mg l(-1)). These calli, after sub-culture on the same medium, produced shoot buds which underwent elongation following transfer of tissues to 6.0 g l(-1) agar-solidified B5 medium containing 30g l(-1) sucrose, IBA (0.01 mg l(-1)) and BAP (1.0 mg l(-1)). Elongated shoots developed roots after transfer to 8.0g l(-1) agar-solidified, hormone-free MS medium with 30 g l(-1) sucrose.
authors.affiliation: Department of Botany, University of Chittagong, 4331, Chittagong, Bangladesh.
authors.forename: S K
authors.initials: SK
authors.lastname: Bhadra
authors.forename: N
authors.initials: N
authors.lastname: Hammatt
authors.forename: J B
authors.initials: JB
authors.lastname: Power
authors.forename: M R
authors.initials: MR
authors.lastname: Davey