Cloning And Characterization Of Fusarium Head Blight Related Genes In Wheat

Fusarium Head Blight Caused by phytopathogenic fungus Fusarium graminearum Schwabe is an important disease of wheat and barley. In recent years, the disease is often outbreak in the yellow river-huai river area and Northwest area of wheat production, resulting in great losses. Fusarium can infect host plants throughout the growth cycle, causing various types of rot phenotype, such as seedling rot, stem rot and ear rot, of which the ear rot cause the most severe damage, sometimes leading to complete yield loses. The fungus produces mycotoxins and secondary metabolites that can contaminate grains of cereal crops, humans and livestock consumed the food made from them can also cause vomiting, diarrhea, dizziness and other acute poisoning and long-term retention in the food chain, during which carcinogens may be produced, so it is a serious threat to human and animal health. Therefore, wheat scab not only reduces grain yield but also edible and industrial value. Research on the molecular mechanism of the interaction between Fusarium graminearum and wheat, and the interaction between pathogenesis-related genes in host cells, exploring disease susceptible or resistance genes, could be of great significance for combating the disease. In this study, using one of xylanases as bait, we screened the wheat cDNA expression library (constructed with wheat cultivars Sumai 3 and Wangshuibai as materials) by yeast two-hybrid (Y2H), an immunophilin like protein was identified, which was used as bait to further screen the library. The main results are as follows:1. FGX, an effector secreted by phytopathogenic fungus Fusarium graminearum was clone by RT-PCR, it was sub-cloned into the pGBKT7 vector and used for screening of cDNA library.2. By screening wheat cDNA expression library constructing with wheat spikes of Wangshuibai and Sumai3 inoculated by Fusarium graminearum, one candidate g ene was screened in the cDNA library of Sumai3, sequencing and bioinformatics an alysis reveal the gene is a putative immunophilin containing FKBP_C superfamily d omain. The open reading frame of the gene consists of 339bp, predicted to encode a protein with 112 amino acids residus, the molecular weight is 12 kDa, so designa ted TaFKBP12(Trticum aestivum FK-506 binding proteins 12 kDa).Two candidate ge ne were screened in the Wangshuibai cDNA library of. One is designated W1, enco ding Rubisco large subunit; the other is designated 67, encoding a pathogenesi-relate d 1(PR-1) like protein. The open reading frame of W1 consists of 1431bp, predicte d to encode a protein consists of 476amino acid residus, the molecular weight is 53 kDa, so designated TaRBC-O53(Trticum aestivum ribulose -1,5- bisphosphatecarboxy lase/oxygenase53kDa). The gene open reading frame of 67 consist of 498bp, predict ed to encode a protein with165 amino acid residus, the molecular weight is 18 kDa, so designated TaSPRP18(Trticum aestivum SCP pathogenesis-related proteins 18 kD a).3. TaFKBP12 was clone by RT-PCR, it was sub-cloned into the pGBKT7 vector and used to screen on the two cDNA librarys4. using TaFKBP12 as bait, we screened the two cDNA librarys, an elicitor response protein containing C2 domain was identified. The open reading frame of the gene consists of 426bp, encoding a protein with 141 amino acid residues, the molecular weight is 15 kDa, so designated TaERGC2d15(Trticum aestivum Excitation response gene 15 kDa)5. semi-quantitative RT-PCR analysis showed that TaERGC2d15 expression in resistant materials of Wangshuibai and Sumai 3, were down regulated after inoculated with Fusarium graminearum.