Plant Regeneration from Protoplasts of Birdsfoot Trefoil, Lotus corniculatus L.

Abstract

A system was established for achieving plant regeneration from callus protoplasts of birdsfoot trefoil. Lotus corniculatus L. A fresh callus was incubated in an enzyme mixture contalnlng 4% Cellulase "Onozuka" RS, 1% Macerozyme R-10, 0.2% Pectolyase Y-23, 0.7M mannitol at pH 5.8 for 3∼4 hr, to liberate the protoplasts. The protoplasts in the modified 8p medium (KAO and MICHAYLUK 1975) divided within 7 days and formed cell clusters and subsequently colonies. The colonies were transferred to the medium of MURASHIGE and SKOOG (1962) supplemented with l-naphthaleneacetic acid (NAA) and kinetin and solidified with agar. The calli thus fomed were again trans-ferred to the medium of MILLER (1963) with different concentrations of indole-3-acetic acid (IAA) and 6-benzyladenine (BA). After 1 month many adventitious buds were formed. The most suitable medium for the adventitious bud formation contained 1.5 mg/l IAA and 1, 5 mg/l BA. These adventitious buds developed to complete plantlets on the medium of NITSCH and NITSCH (1969) Iacking growth regulator.