Genetic recombination by protoplast fusion in Streptomyces (Reprinted fromDevelopments in Industrial Microbiology, vol 21, pg 43-54, 1980)

Genetic recombination by protoplast fusion recently has been demonstrated i n a number of Streptomyces species, and in at least one species of Streptos porangium. Protoplast formation by lysozyme treatment is facilitated by gro wing mycelia in a complex medium containing a partially growth inhibitory l evel of glycine. Protoplast fusion is readily induced by treating protoplas ts with polyethylene glycol, and regeneration of tells from protoplasts can take place on hypertonic media. The efficiency of cell regeneration from p rotoplasts of Streptomyces fradiae and Streptomyces griseofuscus is growth- phase dependent; maximum regeneration is obtained from cells harvested betw een late exponential and stationary growth phases. Recombinant frequencies in intraspecific streptomycete crosses by protoplast fusion are very high a nd typically vary from about 0.5 to 15% of total viable protoplasts. The fr equency of recombinants can be further increased in Streptomyces coelicoior and S. fradiae (and presumably in other Streptomyces) by treating protopla sts with ultraviolet (UV) light. The very high frequencies of recombinants obtained should facilitate construction of superior antibiotic-producing st rains without genetically marking the parental strains. It also should faci litate construction of strains with complex genotypes for intraspecific gen etic mapping studies and may facilitate formation of interspecific streptom ycete hybrids to produce novel antibiotics. Protoplast fusion currently is being used to construct a genetic map of S. fradiae, and preliminary experi ments indicate that the tylA gene, which codes for an enzyme involved in fo rmation of a common intermediate for biosynthesis of the tylosin sugars, is chromosomal.