Technical Research Of Protoplast Fusion Between Pleurotus Eryngii And Pleurotus Pulmonarius

Protoplast fusion breeding technology as one of new cell engineering breedingtechnology is developing rapidly in recent years and it develop a entirely new approach forutilization of wild germplasm, improving working efficiency,shorting the breedingcycle,expanding genetic. Therefore, this technology has respective advantage in ediblefungi breeding area and attracted a wild spread attention. Heat-inactivated protoplastprepared from pl.e0091and protoplast prepared from pl.g0001..1respectivelyprocess were put in practice by the method of adopting the PEG to get one fusant R1-16.Bands specific for both parental strains were observed with fusant R1-16by the methods ofmolecular marker technology, thereby confirming the strain R1-16to be true fusionfinally.The main research results are as follows:1Selection of parent strains. In this study, we select strains pl.e0091and pl.g0001..1asparent strains according to the product of protoplasts and the growth speed of mycelium.2Determining the optimum conditions for the formation, regeneration, and fusionof protoplasts. The optimum conditions of protoplasts formation, regeneration and fusionwere discovered by single factor test and orthogonal test mainly from the following sixaspects:mycelium age, cultivation ways, enzymolysis temperature, kinds of osmolaritystabilizers, kinds of regeneration medium, enzyme concentration. The study showed thatthe process of integration was objective.3Selection of fusant. In this study, we got a genetic stable fusant R1-16by antagonistictest and marks of clamp and heat-inactivated.4Identification of fusion. Antagonistic test demonstrated that there were some geneticdifferences between fusant and its parents. Methods of molecular marker technology suchas RAPD,SRAP,ISSR,SCAR were used to identify fusant and found that the geneticmaterial of fusant is source of the two parents.5Genetic analysis. Genetic marker tests of single spore isolates of R1-16showed thatgenetic material inherited from parents separation occurred during meiosis and isolation ofspecific bands of R1-16were observed.