基于iTRAQ技术筛选甜菜M14单体附加系块根变异性状的相关蛋白

Sugar beet is an important sugar and economical crop in the world, its tuber size and sugar content are two most important agronomic traits. However, few studies on tuber quality regulatory mechanisms of sugar beet have been reported compared to other crops. Sugar beet M14, a monosomic addition line by the intercross of Beta vulgaris L. and B. corolliflora Zoss., contains the B. vulgaris L. genome and chromosome 9 of B. corolliflora Zoss.. The sugar beet M14 has some variated traits compared with its parents, such as smaller taproot, forked tuber, lower sugar content and higher cell lignification, etc. Based on the guide of production demand and main restrictional factors on the development of sugar beet production, our study will perform the investigations on the variable structural features and the dynamic changes of physiological and biochemical processes during taproot development. Then the key stages of root variable development and sugar accumulation in taproot can be understood. To investigate the molecular mechnisms and interrelation on root growth and sugar accumulation, root samples of M14 and B. vulgaris L. in the key stage will be used for iTRAQ LC-MS/MS analysis. The obtained differential expression proteins related to root development and sugar accumulation will contribute to the construction of the molecular regulatory networks on high efficient sugar accumulation in sugar beet. This research will lay the theoretical foundation of culturing new high-sugar varieties, and promote the sustainable development of the country's sugar beet industry.

甜菜是世界上重要的糖料作物和经济作物，其块根大小及含糖量是甜菜最重要的农艺性状，但目前甜菜的块根品质分子调控机制研究仍很滞后。甜菜M14为二倍体栽培甜菜（Beta vulgaris L.）附加野生白花甜菜第9号染色体后获得的单体附加系，其块根表现为块根变小、岔根、糖分降低、高木质化程度等变异性状。本研究以我国甜菜生产重大需求为导向，围绕制约我国甜菜糖业发展的主要因素，研究甜菜M14品系块根变异性状的发育过程及相关生理生化指标动态变化，以确定甜菜块根畸形发育和糖分差异积累的关键时期；采用同位素标记相对和绝对定量（iTRAQ）蛋白质组学技术，筛选关键时期M14品系差异表达蛋白质，并进一步从蛋白质水平探讨甜菜块根发育与糖分积累的相互关系及其分子机理。研究结果将发现与甜菜块根增长与糖分积累相关蛋白质，有助于构建甜菜糖分高效积累的分子调控网络，为培育甜菜高糖新品种，促进我国甜菜产业发展奠定理论基础。

甜菜是世界上重要的糖料作物和经济作物，其块根大小及含糖量是甜菜最重要的农艺性状，但目前甜菜的块根品质分子调控机制研究仍很滞后。本研究以甜菜单体附加系M14品系和二倍体栽培甜菜为材料，通过测定两品种甜菜块根直径的增长曲线确定了水培条件下甜菜块根发育的时间节点，然后采用酚提取法提取了甜菜M14品系及二倍体栽培甜菜在叶丛快速生长期和块根膨大期的总蛋白，经过定量、样品还原、烷基化、胰蛋白酶酶解等步骤，进行 iTRAQ 标记、强阳离子交换分离LC-MS/MS 鉴定。通过Mascot软件共鉴定到5336个与已知图谱相匹配的图谱，匹配到的肽段有2824个，匹配到特有肽段的数量为669个，匹配到蛋白有332个，定量的蛋白有259个。根据变化倍数阈值>1.2或<0.8定义筛选差异表达蛋白质，结果在定量的259个蛋白质中获得了130个差异表达蛋白质，其中甜菜M14品系47个，二倍体栽培甜菜83个，二者共有26个。我们对这些差异蛋白进行了GO分类、互作网络及KEGG代谢通路分析，结果显示在甜菜M14品系中有9个蛋白质定位的两个代谢途径（Amino sugar and nucleotide sugar metabolism和2-Oxocarboxylic acid metabolism）在二倍体栽培甜菜中不存在，二倍体栽培甜菜中有31个蛋白质定位的三个代谢途径（Pyruvate metabolism、Citrate cycle (TCA cycle)和Biosynthesis of secondary metabolites）在M14品系中不存在，这可能是造成二者块根发育存在差异变化的主要原因。该研究结果有助于构建甜菜糖分高效积累的分子调控网络，为培育甜菜高糖新品种，促进我国甜菜产业发展奠定理论基础。