AFLP and STS tagging of Lr19, a gene conferring resistance to leaf rust inwheat

Amplified fragment length polymorphism (A-FLP) markers were used to enrich the map of the wheat chromosomal region containing the Thinopyrum-derived L r19 leaf rust resistance gene. The region closest to Lr19 was targeted thro ugh the use of deletion and recombinant lines of the translocated segment. One of the AFLP bands thus identified was converted into a sequence-tagged- site (STS) marker. This assay generated a 130-bp PCR fragment in all Lr19-c arrying lines tested, except for one deletion mutant, while non-carrier tem plate failed to amplify any product. This sequence represents the first mar ker to map on the distal side of Lr19 on chromosome 7el(1). The conversion process of AFLP fragments to STS markers was technically difficult, mainly because of the presence of contaminating fragments. Various approaches were taken to reduce the frequency of false positives and to identify the corre ct clone. We were able to formulate a general verification strategy prior t o clone sequencing. Various other factors causing problems with converting AFLP bands to an STS assays are also discussed.