稻瘟菌无毒基因的分子检测及其指纹类型分析

针对稻瘟菌中Avr-Pita与AvrPiz-t两个无毒基因,设计特异性引物,利用PCR方法对吉林省103株稻瘟菌菌株进行分子检测,并结合AVR-Pib、AVR-C039、PWL2和ACE1 4个无毒基因在其中85个菌株中的分布情况,绘制出基于这6个无毒基因的指纹类型.结果表明,Avr-Pita基因和AvrPiz-t基因在吉林省稻瘟菌中的分布频率分别为86.41%和62.14%;建立的指纹类型显示,85株稻瘟菌分布于9种类型中,其中类型1为主要类型,其分布频率为40.00%:根据30个已知生理小种的菌株在指纹类型中的分布情况,初步发现类型7与吉林省优势生理小种ZE1可能存在对应关系,并应用于稻瘟菌优势生理小种的特异性分子检测;其他指纹类型与中国生理小种并无明显对应关系.明确了无毒基因Avr-Pita与AvrPiz-t在吉林省稻瘟菌中的分布情况,构建了无毒基因的指纹类型,为水稻抗瘟育种和稻瘟菌优势小种的分子监测提供理论依据.     

Rice ubiquitin-conjugating enzyme OsUBC26 is essential for immunity to the blast fungus Magnaporthe oryzae

The functions of ubiquitin-conjugating enzymes (E2) in plant immunity are not well understood. In this study, OsUBC26, a rice ubiquitin-conjugating enzyme, was characterized in the defence against Magnaporthe oryzae. The expression of OsUBC26 was induced by M. oryzae inoculation and methyl jasmonate treatment. Both RNA interference lines and CRISPR/Cas9 null mutants of OsUBC26 reduced rice resistance to M. oryzae. WRKY45 was down-regulated in OsUBC26 null mutants. In vitro E2 activity assay indicated that OsUBC26 is an active ubiquitin-conjugating enzyme. Yeast two-hybrid assays using OsUBC26 as bait identified the RING-type E3 ligase UCIP2 as an interacting protein. Coimmunoprecipitation assays confirmed the interaction between OsUBC26 and UCIP2. The CRISPR/Cas9 mutants of UCIP2 also showed compromised resistance to M. oryzae. Yeast two-hybrid screening using UCIP2 as bait revealed that APIP6 is a binding partner of UCIP2. Moreover, OsUBC26 working with APIP6 ubiquitinateds AvrPiz-t, an avirulence effector of M. oryzae, and OsUBC26 null mutation impaired the proteasome degradation of AvrPiz-t in rice cells. In summary, OsUBC26 plays important roles in rice disease resistance by regulating WRKY45 expression and working with E3 ligases such as APIP6 to counteract the effector protein AvrPiz-t from M. oryzae.