Screening for bioactive mutants with antitumor activity from an actinomycetic wild-type strain without antitumor activity by antibiotic-resistant mutation technique and by coupled with chemical mutagen-induced mutation

Objective To obtain antibiotic-resistant mutants producing metabolites with antitumor activity from wild-type actinomycete strains without antitumor activity.Methods An actinomycete strain L35-1 was used as an initial strain for obtaining antibiotic-resistant mutants,which is a marine-derived wild-type strain without antitumor activity with an inhibition rate of 2.8% at the 1000 μg/ml of high sample concentration on K562 cells.The antibiotic-resistant mutants both from auto-mutagenesis and chemical mutagen-induced mutagenesis were selected by single colony isolation on antibiotic-containing plates according to the method for obtaining drug-resistant mutants in ribosome engineering.The antitumor activity was assayed by the MTT method using K562 cells for the mutants with aqueous acetone extracts of the whole broth of their fermentation.Results A total of 114 neomycin-resistant (ner) and 68 streptomycin-resistant (str) mutants,all from auto-mutagenesis,was obtained on drug-containing plates.Among them,the 7 ner and 3 str mutants appeared to be bioactive with an inhibition rate above 20% at the 100 μg/ml sample concentration on K562 cells.On the other hand,41 str and 32 ner mutants from DES-induced mutagenesis and 46 ner mutants from NTG-induced mutagenesis were obtained by mutagen-induced mutation coupled with the single colony isolation on antibiotic-containing plates,among which,one str mutant from DES-induced mutagenesis and one ner mutant from NTG-induced mutagenesis were bioactive with an inhibition rate over 20% at the 100 μg/ml sample concentration on K562 cells.Conclusions The present result has revealed that the wild-type actinomycete strains without bioactivity might become a great source initial strains to obtain bioactive mutants by drug-resistant mutation technique.