Developing Molecular Markers And Cloning The Lectin-like Receptor Kinases Gene In Wheat Leaf Rust Near-isogenic Line TcLr45

Leaf rust, caused by Puccinia triticina Eriks., is one of the most severe diseases of common wheat (Triticum aestivum L.) throughout the world, and also the important factor affecting the wheat yield in China. The application of leaf rust resistance genes became the most economical and effective method to control the disease. Wheat leaf rust resistance gene Lr45 derived from the Japnases triticale, until now there is no report has been found about the leaf rust strain overcoming this gene in China, therefore Lr45 is an effective gene and with potential application in China. Related studies were carried out on wheat leaf rust resistance gene Lr45 for developing molecular markers and cloning resistance related gene, which laid a solid foundation for the molecular identification and annotation the resistance mechanism of Lr gene. The major results are as follows:1. 3 SSR markers, 6 RAPD markers and 1 STS marker located on rye 2R chromosome and 4 EST-SSR markers, 31 SSR markers located on wheat 2A chromosome were screened the linkage molecular markers for Lr45. The SSR markers SCM43, SCM75 located on rye chromosome 2R and WMC63 from wheat chromosome 2A amplified the polymorphism between TcLr45 and Thatcher, but with a long genetic distance.2. Wheat leaf rust near-isogenic line TcLr26 and TcLr45 which derived the same heredity background as TcLr25, and susceptible background genotype Thatcher were detected with the SCAR marker Lr25F20/R19 of wheat leaf rust resistance gene Lr25. A unique 1859bp band amplified in TcLr45. The amplification results of 52 wheat leaf rust near-isogenic lines, 6 wheat varieties, 5 rye varieties and 3 triticale varieties validated Lr25F20/R19 is a specific molecular maker with 14cM genetic distance for wheat leaf rust resistance gene Lr45, and named LR45R/F. The results corrected the location of the SCAR marker Lr25F20/R19, and will avoid the unreliable of the marker-assisted selection.3. EST-CAPS was the first time carried on wheat leaf rust near-isogenic line TcLr45 research. 2 of 25 EST markers and 4 of 100 primers/restriction enzyme combinations which located on wheat chromosome 2A short arm C-2AS5-0.78 domain amplified the polymorphism between TcLr45 and Thatcher.4. A pair of pimers was disigned based on the lectin-like receptor kinases gene sequence of wheat leaf rust resistance gene Lr34. A single band was amplified in the cDNA of TcLr45. The fragment was cloned and sequenced, and it is a part of lectin-like receptor kinases gene. Based on the target sequence, the complete sequence of cDNA, 2347bp and encoding 730aa, was obtained through RACE-PCR. SMART software was used to analysis this gene found that it has classic LecRKs structures, and belongs to L type, named as LecRK-LR45.