Cloning And Functional Analysis Of The Melanin Biosynthesis Gene In Setosphaeria Turcica

In order to identify the melanin biosynthetic pathway and the relationship between melanin and the pathogenicity of Setosphaeria turcica, PKS and SCD which belong to the melanin biosynthetic pathway were cloned and the functional analysis were done by bioinformatics analysis, meanwhile analysis of gene-specific inhibitor was completed. It was conclusion that SCD and PKS genes might play an important role in melanin biosynthetic pathway and pathogenicity ofS. turcica.Genemonic DNA and cDNA homologous fragments of the scytalone dehydratase gene were obtained by polymerase chain reaction (PCR) amplification from degenerated primer sets designed on the basis of the conserved amino acid regions of scytalone dehydratase and polyketide synthase domains from others fungis. The completed cDNA sequence of SCD in S. turcica was obtained by the method of SMART-RACE and 3'RACE. There is one open reading frame composed of 181 codons and two deduced introns of 50 and 78 nucleotides in the SCD gene. The deduced amino acid sequence of the SCD showed high similarity to the amino acid sequence of scytalone dehydratase from Bipolaris oryzae. DNA blot analysis indicated that there was a single copy of the SCD gene in the genome.A specific inhibitor--Carpropamid could inhibit the conidial germination and appressorium production of S. turcica within 24h treatment but no evident inhibitory effect after 24h. The experimental results also indicated that E. turcica could not penetrate the surface of corn tissue or increase in the corn tissue.A 4kb gene fragment was obtained by SMART-RACE and Genome Walking. The deduced amino acid sequence of the PKS showed high similarity to the amino acid sequence of polyketide synthase from Cochliobolus heterostrophus. There was the same active motifs as PKS.The research was completed in the genes cloning and the analysis of the gene-specific inhibitor, which layed foundation for making new fungicides.