AFLP And SSR Molecular Markers For Leaf Rust Resistance Gene Lr35 In Wheat

Wheat leaf rust resistance gene Lr35 derived from Aegilops speltoides was transfered to the wheat cultivar Marquis through backcrossing with diploid T. monococcum. The gene was mapped on chromosome 2B. It closely links with the wheat stem rust resistance gene Sr39. Its resistance first appears at the growth stage of two leaves, and completely expresses at six leaves. No virulent Puccinia recondita race was found to Lr35 in the world wheat growth area so far. The complete linkage or tightly linkage markers of Lr35 will be important to isolate and use the gene.Amplified fragments length polymorphism (AFLP) and simple sequence repeat(SSR) analyses were carried out in Thatcher, near-isogenic lines carrying twenty-three different genes conferring resistance against leaf rust and F₂ progeny of TcLr35× Thatcher. The main results were as follows:Most of the 304 pairs primers screened for AFLP analysis provided clear amplification products. Sixty-eight of them amplified polymorphic bands in Thatcher and 23 wheat leaf rust resistance NILs with 22.4 percent of polymorphism. Analysis of 132 F₂ progeny of TcLr35 × Thatcher using sixty-eight primers was carried out and four markers were identified with MAPMAKER 3.0b software that tightly linked to Lr35 resistance trait. Two of the markers were named as P-AGG/M-GGT251 and P-ACC/M-GGA200. The genetic distances between Lr35 and P-AGG/M-GGT₂₅₁ and P-ACC/M-GGA₂₀₀ were 0.8 cM and 2.3 cM respectively. The two DNA fragments were reclaimed from the gels and then amplified. They were cloned and sequenced. The sequences were compared with the known sequences in Genbank with BLAST software. P-ACC/M-GGA200 showed 95% homology to a part of AFLP sequence which linked to fusarium head blight (scab) resistance gene fragment (AF389881). The P-AGG/M-GGT₂₅₁ has an ORF, and it had homology to a hypothetical protein of Oryza sativa.Eighteen pairs of SSR primers were used for polymorphic analysis between TcLr35 and Thatcher. Only one pair of primers named Xgwm501 revealed the polymorphism. From the F₂ progeny of TcLr35× Thatcher, the 108 bp fragment detected by the primer Xgwm501 proved to be a specific DNA fragment linked to Lr35. The genetic distances between the 108 bp fragment and the Lr35 gene were 5.4 cM. No repetitive sequences were found from the fragment.