The conversion of wheat RFLP probes into STS markers via the single-stranded conformation polymorphism technique.
We describe a flexible and general strategy for converting a wheat RFLP-based assay into a PCR-based sequence-tagged site (STS), and have applied it to derive markers for a powdery mildew resistance gene present in a wheat-rye translocation. The concept is based on deriving PCR primers that amplify all of the homoeoloci defined by a single-copy cDNA sequence, and separating the resulting mixture of homoeoamplicons via single-stranded conformation polymorphism (SSCP) gels, which are able to detect minor differences between related DNA sequences. After their separation, the individual homoeoamplicons were sequenced and these were used to define nucleotide polymorphisms that could be exploited to design locus-specific PCR primers. In one case, we were able to demonstrate that the assay was allele specific.
Forsström
Per-Olov
PO
Department of Crop Science, Swedish University of Agricultural Sciences, SE-230 53 Alnarp, Sweden. per-olov.forsstrom@vv.slu.se
Koebner
Robert
R
Merker
Arnulf
A
D001483:Base Sequence; D005819:Genetic Markers; D008969:Molecular Sequence Data; D012150:Polymorphism, Restriction Fragment Length; D018807:Polymorphism, Single-Stranded Conformational; D012434:Secale / Q000235:genetics; D016415:Sequence Alignment; D016324:Sequence Tagged Sites; D014908:Triticum / Q000235:genetics*
D005819:Genetic Markers