氮胁迫条件下稻瘟病菌分泌蛋白与水稻悬浮细胞互作中“活性氧”变化分析  

Changes in Concentration of Active Oxidative Analysis of Rice Blast Secreted Protein and Rice Suspension Cell Interaction under Nitrogen Starvation

作　　者：周晓罡[1,2] 侯思名[3] 苏源[1,2] 陶南[1,2] 丁玉梅[1,2] 姚春馨[1,2] 孙茂林[1,2] 张绍松[1,2] 

机构地区：[1]云南省农业科学院生物技术与种质资源研究所,云南昆明650223 [2]云南省农业生物技术重点实验室,云南昆明650223 [3]昆明大学生命科学与技术学院,云南昆明650031

出　　处：《西南农业学报》2011年第6期2164-2171,共8页Southwest China Journal of Agricultural Sciences

基　　金：国家自然科学基金项目(31060021);云南省自然科学基金项目(2006C0062M;2008ZC100M)

摘　　要：以CH-63和TH-16 2个致病性差异显著的稻瘟病菌株为材料,在氮胁迫培养条件下进行液体培养,经培养2 d后,采用铵盐沉淀及透析纯化提取稻瘟病菌培养滤液中的致病性分泌蛋白。以24个IRRI水稻抗稻瘟病单基因系和普感品种丽江新团黑谷(LTH)的成熟种胚为材料,诱导愈伤组织并构建水稻悬浮细胞系。以构建成功的感病品种LTH和抗病品种IRBL-9的水稻悬浮细胞系为材料,接种稻瘟病菌致病性分泌蛋白并对活性氧浓度变化进行检测。结果表明,感病与抗病品种在接种前12 h均出现活性氧迸发,且均出现2个峰值。比较稻瘟病菌株TH-16和CH-63分泌蛋白接种IRBL-9与LTH后H2O2浓度变化特征,发现分泌蛋白致病性强弱与H2O2浓度变化呈正相关,CH-63产生的强致病性分泌蛋白引起活性氧浓度的快速增加。TH-16产生的弱致病性分泌蛋白接种IRBL-9与LTH后活性氧浓度则增加不明显。上述研究对于深入了解稻瘟病菌与水稻之间的相互识别、信号传导和应答过程,阐明水稻对稻瘟病菌的抗性机制、水稻与稻瘟病菌互作的分子机理具有重要意义。The rice blast fungal strains CH-63 and TH-16 with 2 different pathogenic factors were cultivated in a liquid culture medium under nitrogen starvation.After 2 d of cultivation,the protein secreted by the fungal strains was extracted by ammonium sulfate sedimentation and purified by dialysis.Mature embryos of the above mentioned 24 monogenic rice lines and universally susceptible japonica rice variety Lijiangxintuanheigu（LTH） were used as materials to induce callus in inducing medium I and II;the callus-inducing capability varied among the different rice genotypes,and the callus induction rate ranged from 15 % to 98 %.When the callus was cultivated on a subculture medium for some days,the well-grown calli were isolated to develop cell suspension lines of rice.The shape of the suspension cell was observed during the process of cultivation.At the start of the culture,the cells were abnormal in shape and were long,and their vacuole and cytoplasm were big and rarefied,respectively.At the metaphase of culture,the cells gradually became elliptical,and the vacuole and cytoplasm became small and dense,respectively.The changes in the concentration of active oxidative species in the susceptible variety LTH and resistant variety IRBL-9 suspension cell culture were analyzed after inoculation with the secreted protein obtained from the rice blast fungal strains under nitrogen starvation.These suspension cell cultures produced H2O2 in the first 12 h after inoculation.Two peaks were observed at this time,which indicated differences in H2O2 concentration.A comparison of the concentration of H2O2 produced in the cell suspension culture after inoculation of the secreted protein revealed a positive correlation between the pathogenicity of the secreted protein and H2O2 concentration.H2O2 concentration increased rapidly after stimulation with the secreted protein from the strain CH-63,and IRBL-9 showed faster growth than LTH.In contrast,H2O2 concentration increased slowly after stimulation with the secreted protein from the strain

关 键 词：氮胁迫 分泌蛋白 致病性 悬浮细胞系 活性氧 

分 类 号：S435.111.41[农业科学—农业昆虫与害虫防治]