Preliminary Isolation of Viable Protoplasts from Egg Cells in Facultative Apomictic Guineagrass (Panicum maximum)

Abstract

Preliminary isolation of viable protoplasts from egg cells was carried out using 3 enzyme treatments followed by mechanical manipulation from ovaries of facultative apomictic Panicum maximum. Incubation of ovaries staged at anthesis for up to 6 h allowed the boundary wall of the reproductive cells to become completely digested protoplasts in individual. The egg protoplasts cultured on appropriate medium formed complete cell wall after 2 d of culture, indicating the protoplast's strong viability. The same kinds of ovaries were analyzed with Nomarski differential interference contrast microscopy, indicating sizes and morphologies of the cells corresponded to those of isolated protoplasts. Isolation of egg cell and in vitro fertilization could be useful to clarify the mechanism of reproductive process in apomictic P. maximum.

Establishment of Efficient System for Isolation and Manipulation of Single Protoplasts Containing Aposporous Embryo Sac Initial Cell in Guinea Grass (Panicum maximum)

“…From these results, it is indicated that the protoplasts could be isolated from regional tissues. Here, the 3 kinds of enzyme solutions used in this study were same to that of tomato [12] and that of reproductive protoplasts of apomictic guinea grass [13]. However, the single protoplasts were isolated successfully here only in the use of ovaries at anthesis.…”