木霉菌Tri-milR535跨界靶向番茄Chit4逃避宿主免疫性反应的分子机理研究

木霉菌是一种重要的生防菌，可以定殖植物根系组织内，与宿主植物形成共生体。前期研究发现棘孢木霉与番茄根互作中可以产生一个木霉菌特异性表达的小RNAs:Tri-milR535，它可能通过跨界调控番茄Chit4基因的表达参与两者的互作中，但是目前关于该小RNAs的跨界作用机理尚无人研究。因此，本项目以木霉菌Tri-milR535为研究对象，采用Northern blotting和定量PCR技术明确Tri-milR535表达特征；研究Tri-milR535过表达和沉默突变体菌丝在番茄根皮层定殖行为动态和抗病相关基因的变化；采用烟草共表达体系、5`-RLM-RACE等技术研究Tri-milR535对Chit4基因靶向调控关系；利用Chit4过表达和沉默突变体研究Chit4对木霉菌定殖根皮层能力及对番茄抗病性的影响；揭示Tri-milR535在跨界靶向番茄Chit4基因逃避宿主免疫性反应的分子机理。

Trichoderma is an important biocontrol fungus, which can colonize the root tissue of plant and form symbionts. In previous study, a small RNAs (Tri-milR535) specifically expressed by Trichodermas during the interaction between Trichoderma and tomato roots was found. It may target chitinase gene(Chit4) and regulate its expression. However, the role of Tri-milR535 and its related mechanisms have not been studied yet. In this project, the Tri-milR535 was used as the research objection. Firstly, the expression pattern of Tri-milR535 was analyzed by northern blotting and quantitative PCR. The change of colonization behavior of Trichoderma in root cortex of tomato and related genes expression of resistance disease were studied by the mutants of Tri-milR535 over-expression and Tri-milR535 silence-expression. Secondly, we reveal the targeting relationship between Tri-milR535 and Chit4 using co-expression in tobacco and 5`-RLM-RACE techonology. Then, we analyze the effects of Chit4 on colonizing root cortex and the resistance of tomato by over-expression and silence of Chit4. The results will reveal the molecular mechanism of evasion of host immune response by Tri-milR535 from Trichoderma transboundary target Chit4 of tomato.