The Primary Research Of Sugarcane Pokkah Boeng Disease Pathogen-Fusarium Moniliformis Molecular Biology

sugarcane Pokkah Boeng Disease caused by Fusarium moniliformis is one of common diseases in sugarcane production. In 1921, this disease was reported in Indonesia Java firstly. At present, this disease almost occurs in all the sugarcane production countries and causes a loss(Zhang Mian-guo,2009).This paper researches the preparation and transformation technologies of this disease'Pathogen-Fusarium moniliformis's protoplast. In the paper, the effect of liquid height, centrifugal inertia force, the centrifuge tube size, the rate of acceleration and deceleration, the dip angle of centrifuge tube to the deposition and the regeneration of protoplast are researched. The effect of time of cultured on the regeneration broth without antibiotics to the regeneration rate is researched in the paper too. The type of lyase and the concentration of Lysing Enzyme from Trichoderma harzianum'effect to generating protoplast are introduced in this paper too.In order to understand every gene's function, the author use PCR fragement of TrpC promoter fused with hph ORF to transform the Fusarium moniliformis protoplast, and colned the flanking sequence of the inserted site.Since the MAPK pathway is related with the fungus growth, sexual propagation, conidiation. Serine proteinase is a kind of enzyme that digest protein. These two kinds of genes may relate with molecular pathogenesis. In order to find whether these genes are related with pathogenesis, the author researched these two kinds of genes.In this paper, the MAPK pathway is researched, through homologous recombination, three genes knocked out mutants are got. Research to the knocked out mutant show: the FVEG₀2618.3 knocked out mutant's growth rate was similar with wild strain, the phenotypes of knocked out mutants are similar with wild strain, conidiation doesn't change much, the mutants' pathogenicity is lower than wild strain; the FVEG₀3043.3 knocked out mutants' growth rate is lower than wild strain about one third, the colony color is darker, when the mutant is grown on 0.5M NaCl media, the growth rate increases, but the growth rate is still lower than wild strain, the conidiation doesn't change much, the pathogenicity is similar with wild strain; the FVEG₀4168.3 knocked out mutant's growth rate is similar with wild strain, the phenotype of colony is similar with wild strain, the mutant is very sensitive to osmotic pressure. The mutant can hardly grow on 0.5M NaCl media, the conidiation doesn't change much, the pathogenicity is similar with wild strain.The author also got one serine protease gene FVEG₀0212.3 knocked out mutant. The mutant's growth rate is similar with wild strain, the colony color is redder. The generation and morphology of conidia of the mutant doesn't change much, the pathogenicity is similar with wild strain.