Full-Length CDNA Cloning And Bioinformatic Analysis Of Three Stripe Rust Disease Resistance Related Genes In Wheat

Stripe rust is one of the most important diseases of wheat worldwide. In the past years, the research mainly focused on the cytology and histopathology of the interaction mechanism of wheat and Puccinia striiformis f. sp. tritici. However, less was done on the molecular level. In our laboratory, a suppression subtractive hybridization (SSH) cDNA library of wheat cultivar Suwon11 induced by Puccinia striiformis Chinese race CY23 was constructed. 2,167 ESTs which were subsequently assembled and categorized were obtained with high quality. Herein, three stripe rust resistance related genes, which were picked in the SSH cDNA library, were cloned using RACE (Rapid Amplification of cDNA Ends) technology and in silico cloning method. To further understand the molecules and their function in the interaction of stripe rust fungus and wheat, bioinformatic analysis and expression profile analysis were done.A novel GATA type zinc finger protein gene ZF001 was cloned using RACE technology. ZF001 was 1,973 bp in length and encoded a putative protein with 193 amino acids. Bioinformatic analysis suggested ZF001 had a single zinc finger motif (CX2CX20CX2C) in the C-terminal and a bipartite nuclear targeting sequence near the N-terminal. Molecular evolution analysis demonstrated that ZF001 in wheat might share a common ancestor with other species in plant. In contrast, it had a further relationship with those species in vertebrate based on the conserved zinc finger motif. Expression profile of ZF001 under stress showed that the ZF001 transcripts were up-regulated in the incompatible interaction while down-regulated in the compatible interaction. Taken together, ZF001 might play a key role as a transcription factor in the interaction of stripe rust fungus and wheat.Two genes, which were designated TAOEC01 and TaLSD1 respectively, were cloned using the method of in silico cloning in combination with RT-PCR technology. TAOEC01 encoded a putative oxygen-evolving complex precursor and LSD1 encoded a putative GATA type zinc finger protein.TAOEC01 was 824 bp in length and encoded a putative protein with 197 amino acids. Phylogenetic tree analysis indicated that TAOEC01 might share a common ancestor with other oxygen-evolving complex genes in rice (Oryza sativa) and maize (Zea mays), whereas it had a further relationship with other species. The results of semi-quantitative RT-PCR showed that the transcription profile of TAOEC01 which was up-regulated was similar in the incompatible interaction and the compatible interaction. It was speculated that TAOEC01 was involved in the interaction of stripe rust fungus and wheat by regulating the energy metabolism.TaLSD1, which encoded a polypeptide of 146 amino acids with three conserved motifs (CxxCxRxxLMYxxGASxVxCxxC), was 1, 024bp in length. Phylogenetic tree indicated that TaLSD1 might share a common ancestor with other LSD1 type zinc finger protein genes which contained three conserved LSD1 type zinc finger motifs in rice (Oryza sativa)﹑turnip (Brassica rapa) and Arabidopsis (Arabidopsis thaliana), whereas it had a further relationship with other homologous genes with different number of LSD1 type zinc finger motifs. It was assumed that some information in TaLSD1 had been lost to perform new function. The results of semi-quantitative RT-PCR showed that the transcription profile of TaLSD1, which became normal after being down-regulated in the early stage, was similar in the incompatible interaction and the compatible interaction. It was speculated that the transcription of TaLSD1 was light-induced and TaLSD1 functioned as a negative regulator of programmed cell death (PCD) in the interaction of stripe rust fungus and wheat.