Infecting Structures Observation Of Oidium Heveae On Hydrophobic Surfaces And Semi-quantitative RT-PCR Analysis Of Camp Pathway Genes

Powdery mildew in rubber tree, caused by obligate parasitic fungus Oidium heveae B A. Steinmann, mainly infected young tissue and rubber inflorescence, greatly reduced rubber production while serious harm. However,the study of Oidium heveae focused on observation to pathogen biology and cell biology, very little research have been done on O. heveae about gene function on regulate development of infecting structure.For exploring the molecular regulated mechanism in infection of Oidium heveae, we conducted transcriptome sequencing of rubber powdery mildew interactions during infection, find out the AC and PKA genes of cAMP pathway. Induced Oidium heveae B.A.Steinmann HO-73strain infecting structure development by substratum and analyzed gene expression of cAMP pathway by exogenous treatment, preliminary analyzed of the regulation of cAMP pathway genes for O. heveae infecting structures.The main results as follows:Germination and appressorium formation of12d post inoculation fresh conidium was better than the30d,and leaf age7to10d rubber leaves suitable conidium in detail. Infecting structures development induced by hydrophobic surfaces in Oidium heveae, temperature of germination range22to28℃, temperature of appressorium formation.range22-26℃. Much of secondary hyphae developed in sterile water, and germination and appressorium formation in the nutrient solution of illiquid30%glycerol is slow. Ultraviolet (UV) radiation within10min had no significant effects on germination and appressorium formation of conidium in vitro. Parafilm surface with hydrophobicity is more suitable for inducing rubber powdery mildew infection structure development in vitro.Summarized the process of infecting structure development of0-24hpi under parafilm surface as4morphological developmental stage,such as no germinated (0hpi), germination (4hpi), appressorium formation (6,8hpi) and secondary appressorium (16,24hpi). Isolation total RNA of each development characterized stage by modified Trizol method, the concentration match the transcriptome sequencing requirements.The0hpi,30dpi conidium,24hpi,3dpi infected leaves and healthy leaves of transcriptome libraries were built and sequenced by powdery mildew living culture. Total6910GO annotated genes and270metabolic pathways based on the work in the early stage of the genome. Obtained five AC (adenylate cyclase) and PKA (protein kinase) genes of cAMP pathway by gene prediction and homologous alignment.Detection all cAMP genes expression at the induced time range0-24h, two patterns were significantly different. The expression level of adenylate cyclase (AC) gene was lowest at no germinated, then quickly increased at4,6,8hpi. However, expression level of AC down-regulated at16,24hpi stages which secondary appressorium formatted. The expression level of PKA gene was similar to AC at0-8hpi,but expression level up-regulated at16,24hpi stages followed by the opposite trend of AC gene. Exogenous cAMP and IBMX treatment suggested that O.heveae could formed Appressorium in the non-induced agar surface.