Construction And EST Analysis Of A SSH CDNA Library Of Wheat Leaves Induced By Puccinia Striifomis

Wheat stripe rust, caused by Puccinia striiformis f. sp. Tritici, was one of the most important wheat diseases in China, and with the characteristics of wide distribution, fast prevalence and severe losing. It has been demonstrated that the reasonable use of resistant wheat cultivars is the most effective and economical method to control wheat rust diseases. However, new higher virulent race of P. striiformis often made the original resistant cultivars disability, which had become a prominent question in resistant cultivars application. Therefore, studying on the interaction between wheat and rust fungi can reveal the resistant mechanism of the host plant and the pathogenic mechanism of the rust fungi, which provide further information for selection and reasonable use of resistant wheat cultivars. In the present study, suppression subtractive hybridization (SSH) was adapted to construct a wheat leaf cDNA library induced by P. striiformis race CY23. In order to elucidate the wheat resistant mechanism at the molecular level, global gene expression profile was examined after function annotation and category for qualified expressed sequence tags(ESTs) in SSH cDNA library, and resistance related genes were cloned. Several aspects included in the research were as followed:1 Four methods were used for extracting total RNA from different organs of wheat. After comparison, NaAC/ethanol was an ideal method for extracting total RNA from all detected organs of wheat. The content of immature seed total RNA was the highest among all examined organs in the research. Constitution of wheat leaf rRNA was very complicated, besides of 25S rRNA and 18S rRNA in cytoplasm, there exist large amounts of 23S rRNA, 16S rRNA and 9S rRNA of chloroplast.2 Wheat cultivar shuiyuan11 and P. striiformis race CY23 was used as the initial materials, an incompatible wheat SSH library was successfully constructed with wheat leaves inoculation after 24 h, 48 h and 72 h. There were 2,606 positive clones in SSH library. After extracting plasmid DNA and sequencing, 2,167 qualified ESTs were obtained. 652 unigenes including 333 Contigs and 319 Singlets were acquired after clustering by CAP3 software. Results of function annotation and category showed: in those function known unigenes, the...