甘蔗响应甘蔗鞭黑粉菌侵染的生理和分子基础研究

In China, the main threat to the production of sugarcane is sugarcane smut caused by Sporisorium scitamineum. Due to its special pathogenic process, the molecular mechanism of how to infect the host is still unclear. Furthermore, there is no effective way to prevent this disease. The applicants' former research showed that the b locus gene plays a key role in the pathogenic process of sugarcane Sporisorium scitamineum. In this project, the mutant strain SsΔMAT-1b with b locus gene deletion will be used, the main research is focused on the interaction between the strain ROC22 with high smut susceptibility and Sporisorium scitamineum, in order to find out the differentially expressed susceptibility genes and proteins in sugarcane induced by b locus gene. through using LSCM and qRT-PCR to detect the infection and colonization process of mating strain Ss△MAT-1b-GFP and MAT-2-RFP, which were labeled with green fluorescence (GFP) and red fluorescence (RFP), respectively, in order to get the colonization model in sugarcane; by measuring the effects of Sporisorium scitamineum infection on the physiological and biochemical metabolism of sugarcane, and combined with the correlation analysis of transcriptome and proteome data, in order to identify the key genes in response to the infection of Sporisorium scitamineum, analyse the relation among inter-genetic metabolic pathways. Finally, this project will provide the basic information for the development of sugarcane smut preventive technologies and the sugarcane variety improvement.

由甘蔗鞭黑粉菌引起的甘蔗黑穗病是威胁我国甘蔗安全生产最主要的病害。由于其致病过程特殊，目前其侵染寄主的分子机理仍不清楚，亦无有效的防治方法。申请人团队研究证实b位点基因在甘蔗鞭黑粉菌致病过程中起关键的作用。本项目拟选用b位点基因缺失的突变体菌株Ss△MAT-1b，以其与高感黑穗病甘蔗品种ROC22的互作为主要研究内容，找出甘蔗受b位点基因诱导差异表达的感病性基因和蛋白；用激光共聚焦显微镜和qRT-PCR技术监测分别用绿色荧光（GFP）和红色荧光（RFP）标记的交配型菌株Ss△MAT-1b-GFP和MAT-2-RFP侵染定殖进程获得其在甘蔗体内的定殖组织模式；通过测定黑粉菌侵染对甘蔗相关生理生化代谢的影响，结合转录组学（RNA-Seq）和蛋白质组学（iTRAQ）数据关联分析，挖掘出甘蔗响应黑粉菌侵染的关键感病性基因，分析基因间代谢通路关系，为甘蔗黑穗病防控技术开发和甘蔗品种改良提供基础信息。

由甘蔗鞭黑粉菌引起的甘蔗黑穗病是威胁我国甘蔗安全生产最主要的病害。本研究在证实b位点基因在甘蔗鞭黑粉菌致病过程中起关键的作用的基础上，选用b位点基因缺失的低毒力突变体菌株Ss△MAT-1b和高毒力野生型菌株分别接种高感黑穗病甘蔗品种GT11-80蔗芽，发现黑粉菌致病关键b位点基因缺失后，甘蔗的β-1,3-葡聚糖酶、苯丙氨酸解氨酶、过氧化氢酶、过氧化物酶、几丁质酶、赤霉素含量升高；而超氧化物歧化酶、脱落酸、生长素、超氧阴离子自由基、过氧化氢含量降低。利用转录组学（RNA-Seq）和蛋白质组学（iTRAQ）技术测序并对获得的差异表达基因和差异表达蛋白进行功能分析，发现野生型高毒力菌株侵染甘蔗时，能被甘蔗在短时间内识别并刺激甘蔗做出一系列的生理反应，并影响甘蔗的光合作用等生理活动；当b位点基因缺失后，黑粉菌的定殖及对甘蔗免疫系统的激活等受到严重影响；黑粉菌野生型菌株主要通过影响甘蔗的叶绿体、光合作用以及核糖体的代谢活动等对甘蔗产生为害。研究结果挖掘出甘蔗响应黑粉菌侵染的潜在关键感病性基因，进一步揭示了甘蔗响应黑粉菌侵染的生理和分子机理。