New polyketides from chemical mutant strain of sponge-associated fungus emericella variecolor xsa-07-2 / 中国药学杂志

OBJECTIVE: To study the new bioactive secondary metabolites of diethyl sulfate chemical mutant strain of sponge-associated fungus Emericella variecolor XSA-07-2. METHODS: Diethyl sulfate was used to make chemical mutagenesis of strain XSA-07-2, and one mutant strain M8 was chosen for large-scale fermentation to generate new secondary metabolites. The compounds were isolated and purified by chromatography on silica gel and ODS reversed-phase column and semi-preparative HPLC techniques. And their structures were identified by their physicochemical properties and NMR, MS data analysis. RESULTS: Three new polyketides 1-3 were isolated from the extract of the solid fermentation culture of mutant strain M8. Compound 3 showed moderated antioxidant activity with IC50 of (13.58±0.14) μg·mL-1 by DPPH assay. CONCLUSION: Diethyl sulfate chemical mutagenesis can stimulate sponge-associated fungus Emericella variecolor XSA-07-2 mutant strain M8 to produce new antioxidant polyketides.

BSA and molecular markers screening for salt stress tolerant mutant of Petunia obtained in in vitro culture / Rastreamento de BSA e marcadores moleculares para mutante tolerante ao estresse salino de Petunia obtido em cultura in vitro

ABSTRACT: In this study, we performed BSA to identify genetic markers linked to salt tolerance. We tested the genetic diversity among four bulked DNA samples of EMS induced mutant clones and one bulked DNA sample of non-mutated clone of Petunia for salt tolerance in in vitro callus cultures using RAPD and ISSR markers. Out of the 36 RAPD and 16 ISSR primers identified, 25 and 13 were effectively used to amplify genomic DNA of all the five bulked samples, respectively. In total, 114 RAPD amplifications products were obtained, of which 28% were polymorphic and 2% were genotype-specific bands. Out of the 64 ISSR amplification products obtained, 51% were polymorphic and 1% was genotype-specific bands. Results of this study indicated the existence of two patterns of distorted segregation among the studied markers. The first one indicates the differences between non-mutated clones of Petunia and its putative mutants. The second one was observed only between putative mutants and putative mutants tested for salt tolerance in in vitro culture. Both RAPD and ISSR analysis successfully detected the association with changes induced by chemical mutagenesis and salinity. Furthermore, our results indicate that BSA method can be useful in the rapid detection of molecular markers for further marker-assisted selection.

RESUMO: Neste estudo, realizamos BSA para identificar marcadores genéticos ligados à tolerância ao sal. Testamos a diversidade genética entre quatro amostras de DNA volumoso de clones mutantes induzidos por EMS, e uma amostra de DNA volumoso de clone não mutado de Petunia para tolerância a sal em culturas de calos in vitro usando marcadores RAPD e ISSR. Dos 36 primers RAPD e 16 ISSR identificados, 25 e 13 foram efetivamente usados para amplificar o DNA genômico de todas as cinco amostras, respectivamente. No total, foram obtidos 114 produtos de amplificação RAPD, dos quais 28% eram polimórficos e 2% eram bandas específicas de genótipos. Dos 64 produtos de amplificação ISSR obtidos, 51% eram polimórficos e 1% eram bandas específicas de genótipo. Os resultados deste estudo indicam a existência de dois padrões de segregação distorcida entre os marcadores estudados. O primeiro indica as diferenças entre os clones não mutantes de Petúnia e seus mutantes putativos. O segundo foi observado apenas entre mutantes putativos e mutantes putativos testados quanto à tolerância ao sal em cultura in vitro. Tanto a análise RAPD quanto a ISSR detectaram com sucesso a associação com alterações induzidas por mutagênese química e salinidade. Além disso, nossos resultados indicam que o método BSA pode ser útil na detecção rápida de marcadores moleculares para posterior seleção assistida por marcadores.

Generation and Evaluation of High beta-Glucan Producing Mutant Strains of Sparassis crispa

A chemical mutagenesis technique was employed for development of mutant strains of Sparassis crispa targeting the shortened cultivation time and the high beta-glucan content. The homogenized mycelial fragments of S. crispa IUM4010 strain were treated with 0.2 vol% methyl methanesulfonate, an alkylating agent, yielding 199 mutant strains. Subsequent screening in terms of growth and beta-glucan content yielded two mutant strains, B4 and S7. Both mutants exhibited a significant increase in beta-glucan productivity by producing 0.254 and 0.236 mg soluble beta-glucan/mg dry cell weight for the B4 and S7 strains, respectively, whereas the wild type strain produced 0.102 mg soluble beta-glucan/mg dry cell weight. The results demonstrate the usefulness of chemical mutagenesis for generation of mutant mushroom strains.

Breeding of New Strains of Mushroom by Basidiospore Chemical Mutagenesis

Chemical mutagenesis of basidiospores of Hypsizygus marmoreus generated new mushroom strains. The basidospores were treated with methanesulfonate methylester, an alkylating agent, to yield 400 mutant monokaryotic mycelia. Twenty fast-growing mycelia were selected and mated each other by hyphal fusion. Fifty out of the 190 matings were successful (mating rate of 26.3%), judged by the formation of clamp connections. The mutant dikaryons were cultivated to investigate their morphological and cultivation characteristics. Mutant strains No. 3 and No. 5 showed 10% and 6% increase in fruiting body production, respectively. Eight mutant strains showed delayed and reduced primordia formation, resulting in the reduced production yield with prolonged cultivation period. The number of the fruiting bodies of mutant No. 31, which displayed reduced primordial formation, was only 15, compared to the parental number of 65. Another interesting phenotype was a fruiting body with a flattened stipe and pileus. Dikaryons generated by mating with the mutant spore No. 14 produced flat fruiting bodies. Further molecular biological studies will provide details of the mechanism. This work shows that the chemical mutagenesis approach is highly utilizable in the development of mushroom strains as well as in the generation of resources for molecular genetic studies.