The invention relates to the field of
microbial genetic transformation, and discloses a preparation method of Ilyonectria robus
fungus protoplast and a construction method of a genetic transformation
system.The preparation method comprises the following steps that S11, I.robus hyphae are prepared, after I.robus is cultured on a PDA
solid culture medium for 14 days, the I.robus hyphae are inoculated to a CMC
liquid culture medium to be subjected to shake-flask culture, and spores are filtered and collected; diluting the
spore concentration, inoculating into a YEPD
liquid culture medium, carrying out shake-flask culture, filtering and collecting hyphae; and S12, preparing enzymatic
hydrolysate, namely adding the hyphae prepared in the step S11 into the enzymatic
hydrolysate prepared in the step S12, carrying out
shake flask enzymolysis, filtering, centrifuging filtrate, discarding supernate, adding a 0.7 M NaCl solution, centrifuging, and discarding the supernate, so as to obtain the I.robus
fungus protoplast. The protoplasts with proper quantity and good quality can be efficiently obtained, and the
transformation efficiency of a subsequent genetic transformation
system is improved; and the constructed genetic transformation
system can be used for construction of a
mutant library of the I.robusta strain, interaction of a host
plant and the I.robusta strain and research on the
pathogenesis of the I.robusta strain.