Isolation of genes expressed during compatible interactions between leaf rust (Puccinia triticina) and wheat using cDNA-AFLP

Abstract

The rust fungi are obligate biotrophic pathogens that depend on living host tissue for their growth. In compatible interactions they go through a number of developmental stages to form intercellular hyphae and haustoria within host cells, through which they obtain their nutrients. Here we have exploited the cDNA-AFLP technique to isolate fragments of wheat and rust genes that are expressed at specific defined time-points during the infection process. A number of these sequences were used as probes in Northern hybridizations and in RT-PCR to confirm their expression patterns, and were also characterized by PCR analysis and Southern hybridizations to determine whether they are of fungal or wheat origin. A cDNA library was constructed from pooled RNAs extracted from days 5 and 7 after inoculation, and this library was screened to isolate full-length cDNAs of selected sequences. Sequence analysis of these cDNA fragments and clones revealed similarities amongst the fungal genes to a chitinase, a sorbitol utilization protein, an arabinitol dehydrogenase and a proteasome regulatory unit, whilst in wheat, we identified sequences with homology to a katanin and a cell enlargement protein.