Isolation, Identification And Toxin Detection Of Fusarium Graminearum And The Expression Of Gene Tri101 Of Degrading Toxin

Wheat scab, also called red head wheat, is caused by Fusarium Graminearum. It is a destructive disease that can seriously influence the yield of wheat in the wheat growth area. The yield loss may reach 50% in some areas. Wheat scab is distributed in damp and semi-humid areas, especially in humid temperate regions. In Chin, Wheat scab is distributed mainly in the Yangtze River basin and coastal region, especially in Jiangsu Province. When wheat is infected by F. Graminearum, dozens of mycotoxins are produced in diseased seeds, of which, three kinds of toxin, Fumonisin(FB1), Deoxynivalenol (DON) and Zearalenone (ZEN) are most important. Toxin produced by F. Graminearum can brings high risk to human and animal life, therefore, it is very important to study how to degrade these toxins by using biological measures.In this paper, the technology of HPLC was used to analysis the toxins in 100 varieties of wheat from Jiangsu province. It showed that DON is existed in all of the varieties, and most of varieties were contaminated with ZEN. The most serious contamination occurred in variety 3210, in which, detection quantity of DON was 2060.00μg/L, and that of ZEN was 3048.88μg/L. A strain that has high toxin-producing ability was successfully isolated from variety 3210 by using molecular identification. There is a very high phylogenetic relationship between the strain (F3210) and F15. Tri101, a gene for degradeing toxin was cloned from the the DNA of strain F3210. It was reported that the gene was a structural genes encoding the trichothecene 3-O-acetyltransferase, and it was related to resistance of Fusarium to toxin. The prokaryotic and eukaryotic expression systems are successfully constructed to express the target proteins. The results showed that, after 3 hours induction with IPTG, the recombinant strain BL21 (DE3)-pET29a-Tri101 had the highest expression quantities, and the expression products existed in the form of inclusion bodies. The optimal imidazole concentration was 200mM in the process of purifying expression products using nickel column. Expression proteins was also produced in the supernatant when the recombinant strain X33-pPICZa-A-Tri101 was induced by 0.5% methanol. Further work will be modifying the expression vector to obtain the active protein, the work will lay foundation for further studying the role of Tri101 and the host resistance of wheat scab.