Physical mapping of wheat-Aegilops longissima breakpoints in mildew-resistant recombinant lines using FISH with highly repeated and low-copy DNA probes

Fluorescence in situ hybridization (FISH) with multiple probes, consisting of highly repeated DNA sequences (pSc119.2 and pAs1) and of a low-copy, 3BS -specific RFLP sequence (PSR907), enabled determination of the physical pos ition of the wheat-alien breakpoints (BPs) along the 3BS and 3DS arms of co mmon wheat recombinant lines. These lines harbour 3S(1)S Aegilops longissim a segments containing the powdery mildew resistance gene Pm13. In all 3B re combinants, the wheat-Aegilops longissima physical BPs lie within the inter val separating the two most distal of the three pSc119.2 3BS sites. In all such recombinants a telomeric segment, containing the most distal of the pS c119.2 3BS sites, was in fact replaced by a homoeologous Ae. longissima seg ment, marked by characteristic pSc119.2 hybridization sites. Employment of the PSR907 RFLP probe as a FISH marker allowed to resolve further the criti cal region in the various 3B recombinant lines. Three of them, like the con trol common wheat, exhibited between the two most distal pSc119.2 sites a s ingle PSR907 FISH site, which was missing in a fourth recombinant line. The amount of alien chromatin can thus be estimated to represent around 20% of the recombinant arm in the three former lines and a maximum of 27% in the latter. A similar physical length was calculated for the alien segment cont ained in three 3D recombinants, all characterized by the presence of the Ae . longissima pSc119.2 sites distal to the nearly telomeric pAs1 sites of no rmal 3DS. Comparison between the FISH-based maps and previously developed R FLP maps of the 3BS-3S(1)S and 3DS-3S(1)S arms revealed substantial differe nces between physical and genetic map positions of the wheat-alien BPs and of molecular markers associated with the critical chromosomal portions.