Isolation of biochemical mutants using haploid mesophyll protoplasts of Hyoscyamus muticus : I. A NO 3 (-) non-utilizing clone.

A population of 3070 clones derived from N-methyl-N'-nitro-N-nitrosoguanidine (MNNG)-treated mesophyll protoplasts of haploid Hyoscyamus muticus was tested for amino-acid auxotrophy without enrichment. One clone (MA-2) was stably and specifically dependent on casein hydrolysate and could be fed also by a number of single amino acids or by other reduced nitrogen sources. MA-2 was found to be chlorate resistant and devoid of in vivo nitrate reductase activity under inductive conditions. Permissive and restrictive growth conditions for MA-2 were investigated more closely and media were found promoting morphogenesis. Selection and testing of clones were complicated by an unspecific growth stimulation of some wild type cultures by amino acids, thiamine and m-inositol.