Gy. Chen et al., CULTURE AND REGENERATION OF PROTOPLASTS FROM SHOOTS OF ASPARAGUS CULTURES, International journal of plant sciences, 158(5), 1997, pp. 543-551
A reliable culture and regeneration protocol has been established for
asparagus (Asparagus officinalis L.) protoplasts isolated from shoots
of in vitro plants. Studies Focused on factors to maximize plating eff
iciency, colony formation, and plant regeneration. The optimized proto
col involved the culture of asparagus protoplasts embedded in agarose
beads through a series of defined media. Etiolated shoot cultures were
a superior source for protoplasts than green shoot cultures, and grow
ing feeder cells were critical for high plating efficiency and colony
formation. In the presence of growing asparagus feeder cells, asparagu
s protoplasts initiated cell divisions within 2 d of plating in agaros
e with plating efficiencies up to 20%. About 80% of the protoplasts in
itiating cell divisions developed into colonies of at least 25-30 cell
s, of which 25% initiated somatic embryos. Complete plants were regene
rated from the embryos at a frequency over 20%. Overall, ca. 0.8% of t
he initially isolated protoplasts regenerated into complete plants wit
h this protocol. Plants regenerated from protoplasts have been transfe
rred to soil and maintained phenotypically normal morphology for over
2 yr. Chromosome counts confirmed the diploid status (2n=20) of plants
regenerated by somatic embryogenesis.