摘要
【目的】为了研究柑橘衰退病毒(Citrus tristeza virus,CTV)诱导甜橙差异表达蛋白的分离和鉴定,【方法】以接种了CTV的甜橙植株为实验组,健康植株为对照组,利用双向荧光差异凝胶电泳(2D-DIGE)技术分析CTV诱导的甜橙差异表达蛋白。【结果】以t检验P<0.05和相对表达量≥1.5的水平作为判别标准,获得了91个CTV诱导的差异表达蛋白点,在实验组中含量高于对照组的蛋白56个,含量低于对照组的蛋白35个。通过MALDI-TOF质谱分析成功鉴定从中选择的37个蛋白点,其中19个蛋白功能明确,16个为预测或假定具有某种蛋白功能,包括与抗氧化相关的硫氧还蛋白、铁氧还蛋白、超氧化物歧化酶和抗坏血酸过氧化酶,与代谢相关的磷酸甘油酸酯水解酶、木葡聚糖内糖基转移酶、变味蛋白,分子伴侣热激蛋白,折叠酶肽酰脯氨酰顺反式异构酶,以及与光合代谢相关的蛋白等。【结论】CTV的侵染影响到甜橙各种复杂的生物学过程。
【Objective】To determine the probability of 2D-DIGE technology to identify the differentially expressed proteins in sweet oranges induced by Citrus tristeza virus,CTV.【Method】 The same weight leaf samples of each plant were mixed from CTV-inoculated sweet orange plants and healthy plants,respectively.Total proteins were extracted from above mixed samples with three replications.Each protein sample was labeled with three different CyDyes Cy2,Cy3 and Cy5,and Cy2-labeled sample was used as an internal standard pooled from all the samples.Labeled protein samples were separated with 2-D DIGE and differential protein spots were picked out.MALDI-TOF-MS and bioinformatics were adopted to identify and interpret the significance of differentially expressed proteins.【Result】Total 91 Differential protein spots were detected with statistical variance of two groups（relative average volume ratio ≥1.5;t-test,P0.05）.Among these proteins,56 protein volumes of CTV group were higher than that of healthy group,and 35 proteins lower.PMFs of 37 proteins were obtained by MAIDI-TOF-MS analysis.Through searching NCBI,19 protein functions were clear,16 proteins predicted or putative,and 2 proteins unknown.The identified proteins were involved mainly in metabolism（including 2-Phospho-D-glycerate hydrolase,NADP-isocitrate dehydrogenase,Xyloglucan endotransglycosylase,Adenylate kinase）,antioxidant activity（including Thioredoxin H-type 5,Ferredoxin-NADP reductase,L-ascorbate peroxidase T,Iron superoxide dismutase）,photosynthesis（including Ribulose 1,5-bisphosphate carboxylase/oxygenase large or small subunit,Oxygen-evolving enhancer protein 1 and 2,Chlorophyll a/b-binding protein precursor） and molecular chaperone（such as Heat shock protein）.【Conclusion】 2D-DIGE can be used to identify the differentially expressed proteins in sweet oranges induced by different pathologic CTV strains in order to clarify the mechanism of CTV pathogenesis.
出处
《果树学报》
CAS
CSCD
北大核心
2013年第1期16-21,I0002,共7页
Journal of Fruit Science
基金
国家自然科学基金(30771485)
长江学者和创新团队发展计划项目(PCSIRT
IRT0976)
公益性行业(农业)科研专项(201203076)
关键词
柑橘
衰退病毒
蛋白
双向荧光差异凝胶电泳
Citrus
Citrus tristeza virus
CTV
Protein
Fluorescence two-dimensional difference gel electrophoresis
