Nuclear Transplantation into Plant Protoplasts:Optimal Conditions for Induction and Determination of Nuclear Uptake

Abstract

Summary A simple method, based on differential staining of the nuclei, has been developed for an accuratequantitative estimation of the frequencies of uptake of nuclei by plant protoplasts. Foreign nuclei, isolated from cell suspension cultures of Brassica nigra, Glycine max, Vicia hajastana , and Datura innoxia were introduced into the protoplasts of two auxotrophic mutants of Datura innoxia using the uptake inducing solution which contained glycine, polyethylene glycol and calcium chloride. The factors which affected the frequency of nuclear uptake were the type and level of polyethylene glycol, the pH of the uptake inducing solution and the period of incubation. Addition of dimethylsulfoxide to the uptake inducing solution also increased the uptake frequency. Under optimized conditions the frequencies of uptake of the nuclei, isolated from four different cell cultures, by both the receptor protoplast systems varied between 4 - 6%.