ARS | Publication request: Application of complementation tests in identifying pathogenicity determinants of the chickpea pathogen Ascochyta rabiei

Submitted to: Phytopathology
Publication Type: Abstract Only
Publication Acceptance Date: April 20, 2009
Publication Date: July 17, 2009
Citation: White, D., Chen, W. 2009. Application of complementation tests in identifying pathogenicity determinants of the chickpea pathogen Ascochyta rabiei.Phytopathology. 99:S141

Technical Abstract: The necrotrophic pathogen Ascochyta rabiei causes chickpea Ascochyta blight. Very little is known about its pathogenicity mechanisms. The objective of this research was to identify pathogenicity determinants of A. rabiei using complementation tests. The hygromycin-resistant mutant ArW519 was non-pathogenic on chickpea by virtue of a single T-DNA insertion. Genomic DNA regions corresponding to the T-DNA insertion site were isolated from a phage library of a wild type strain AR628. Six genomic fragments were moved separately into a T-DNA vector with geneticin resistance and used to re-transform the mutant ArW519. The pathogenicity of eight double transformants (resistant to both antibiotics) recovered from independent T-DNA integration events was compared to that of the parent mutant ArW519 and of the wild-type AR628 on chickpea using a minidome bioassay. One of the six clones tested was able to functionally restore pathogenicity to the ArW519 mutant. Each of the two antibiotic cassettes in the double transformants was verified to be intact. The complementing genomic fragment contains about 3000 bp of DNA upstream of the T-DNA insertion site and about 1000 bp of DNA downstream, and it carries DNA sequences that are similar to retrotransposon Molly from Stagonospora nodorum and the AvrLM1 avirulence gene from Leptosphaeria maculans.