Two dominant sequence characterized amplified region (SCAR) markers (linked at 3.0 cM, coupling phase) were constructed for the strawberry (Fragaria ×ananassa Duch.) gene Rpf1. This gene confers resistance to red stele root rot, caused by the soil-born fungus Phytophthora fragariae Hickman var. fragariae. The SCAR markers were developed originally from the sequence of RAPD OPO-16C(438) that is linked in repulsion phase to the Rpf1 allele. This SCAR primer set produced multiple bands in the resistant test progeny and in some of the susceptible progeny; therefore, new SCARs were developed based on the sequence differences among these bands. These new SCARs were linked in coupling phase to the Rpf allele and mapped to the same location as the original RAPD OPO-16C(438). The SCAR markers, as well as some additional RAPD markers known to be linked to Rpf1, were shown to be highly conserved in linkage to the gene based on examination of 133 European and North American Fragaria L. sp. cultivars and breeding selections. These flanking RAPD and SCAR-PCR markers can be used in breeding programs for the selection of red stele (Rpf1) resistance.