The Screening And Characterization Of Botrytis Cinerea Mutants Generated By T-DNA Insertion

The transformant BCt41 was found by screening the transformants of Botrytis cinerea produced by Agrobacterium tumefaciens-mediated method, which lost the ability of producing sclerotia, and significantly reduced spores production on PDA medium, but showed the stronger pathogenicity to tomato leaves and fruits compared to the wild type strain. PCR amplification and Southern blotting hybridization on the genomic DNA of BCt41 showed that the genomic DNA of BCt41had carried steady heritable single copy T-DNA. The 264 bp flanking sequence of T-DNA insertion sites was gained by amplifing genomic DNA of BCt41 using thermal asymmetric interlaced PCR (TAIL-PCR) technique. Comparison of 264 bp sequence with Botrytis cinerea database showed that T-DNA insertion sites located in exon 2 of the gene BC1G₀2176.1. The 1206 bp coding region of BC1G₀2176.1 gene with a 54 bp intron, encoded 383 amino acid, and the function of BC1G₀2176.1 gene was unknown. Our results lay foundations for further studying BC1G₀2176.1 gene functions in Botrytis cinerea involving the development of conidia and the formation of sclerotia as well as pathogenicity to tomato leaves and fruits.