Investigation of pathogenicity determinants in a hemibiotrophic ascomycete  Leptosphaeria maculans
L. BURKETOVA (1), M. Novakova (1), D. Stehlik (2) (1) Institute of Experimental Botany, Academy of Sciences of the Czech Republic, Czech Republic; (2) University of Chemistry and Technology Prague, Faculty of Food and Biochemical Technology, Czech Republic

Agrobacterium tumefaciens-mediated random insertional mutagenesis (ATMT) is comm used to investigate pathogenicity determinants in pathogenic fungi.  Analysis of Leptosphaeria maculans mutants with a random insertion containing a GFP tag identified a F2-4 mutant exhibiting a remarkable hypovirulent phenotype. On susceptible Brassica napus cultivars Columbus and Eurol about 50 % less necrotic lesion area was observed for F2-4 isolate in comparison to wild-type isolate NzT4. We used TAIL-PCR (thermal asymmetric interlaced polymerase chain reaction), a powerful tool for the recovery of DNA fragments adjacent to known sequences, to map the position of the T-DNA and to identify surrounding genes that might be affected in their expression. The T-DNA fragment from A. tumefaciens was found to be located in between two L. maculans genes. We performed an expression analysis of these and surrounding genes in in vitro cultivated mycelia of F2-4 and wild-type NzT4 isolates using RT-qPCR. The expression of the two adjacent genes and also of one additional neighboring gene. The expression of the both genes located on the left from the insertion was suppressed. Therefore, the observed decrease in pathogenicity could be related to one of these genes. We intend to elucidate which of the genes is/are responsible for a hypovirulent phenotype of the F2-4 isolate using a post-transcriptional gene silencing. This work was supported by the grant 13-26798S from the Czech Science Foundation.

Abstract Number: P9-243
Session Type: Poster