Genome-wide isolation of resistance gene analogs in maize (Zea mays L.)

Conserved domains or motifs shared by most known resistance (R) genes have been extensively exploited to identify unknown R-gene analogs (RGAs). In an attempt to isolate all potential RGAs from the maize genome, we adopted the following three methods: modified amplified fragment length polymorphism (AFLP), modified rapid amplification of cDNA ends (RACE), and data mining. The first two methods involved PCR-based isolations of RGAs with degenerate primers designed based on the conserved NBS domain; while the third method involved mining of RGAs from the maize EST database using full-length R-gene sequences. A total of 23 and 12 RGAs were obtained from the modified AFLP and RACE methods, respectively; while, as many as 109 unigenes and 77 singletons with high homology to known R-genes were recovered via data-mining. Moreover, R-gene-like ESTs (or RGAs) identified from the data-mining method could cover all RACE-derived RGAs and nearly half AFLP-derived RGAs. Totally, the three methods resulted in 199 non-redundant RGAs. Of them, at least 186 were derived from putative expressed R-genes. RGA-tagged markers were developed for 55 unique RGAs, including 16 STS and 39 CAPS markers.