Mutagenesis Of Astragalus Variabills Bunge Endophytic Fungi And Screening Of SW Mutant Strains

Locoweeds are toxic plants of the genera Astragalus and Oxytropis containing swainsonine.Swainsonine is the principal agent responsible for locoism in animals.Recently,the fungal endophytes are isolated frequently from locoweeds,Undifilum spp,are believed that responsible for the production of swanisonine in locoweeds.If the swainsonine production in Undifilum spp is inhibited,the locoweed's toxicity would be reduced or eliminate.Currently,the mechanism was few known about the biosynthesis of swainsonine in Locoweed's endophytic fungi.The research about influence of different factors on swainsonine production will be benefit for building the important basis for further to elucidate the swainsonine biosynthetic pathways and molecular regulation mechanism.The principal purpose of this paper was to isolate and identify the swainsonine-producing fungal endophytes,U.oxytropis,from locoweeds in Ningxia.The influence of different factors on swainsonine production was estimated in U.oxytropis.The protoplasts of U.oxytropis was developed and regenerated This study will provide a important science basis for further to elucidate the swainsonine biosynthetic pathways and build a non-toxic fungus.1.The results indicated that three isolates are harvested from Astragalus variabills Bunge.All of the strains were determined to synthesize swainsonine.The morphology and genetic evolution relationship of the strain was closely related to the Undifilum oxytropis.Therefore,the strain was identified as U.oxytropis in this study.2.The highest production rate of protoplast was obtained when NM-UA003 was cultured in the liquid medium for 15 d at 22 ?,and then the mycelia were digested by a compound enzyme consisting of 0.6 mol/L MSO4,1%cellulase,1%lywallzyme and 1.2%snailase for 2 h at room temperature,90 r/min.As for protoplast regeneration,a digestion time of about 2 h was preferable.The protoplast concentration can be achieved at 8 × 107?1.2 × 108 mL.The protoplasts of endophytic fungi were transformed in vitro by using fungal expression vectors pAN7,pDL2 and pFL2,and could be grown on the regeneration medium after transformation.3.The mutants were mutagenized by UV and EMS,NTG mutagens,and 79 mutants were randomly selected for enrichment and culture.The results of SW analysis showed that six mutants with SW mutations were significantly changed by UV mutagenesis(P<0.05).When SW mutagenesis was used,12 strains of SW were significantly changed(P<0.05).When mutagenic treatment was used for NTG,8 mutants with significant changes in SW yield were obtained(P<0.05).