中国农业科学 ›› 2014, Vol. 47 ›› Issue (16): 3121-3131.doi: 10.3864/j.issn.0578-1752.2014.16.001

• 作物遗传育种·种质资源·分子遗传学 •    下一篇

耐热和热敏感水稻应答灌浆初期高温胁迫过程中的差异表达蛋白质鉴定

廖江林1, 2, 宋宇2, 钟平安2, 周会汶2, 张宏玉2, 黄英金1, 2   

  1. 1、江西农业大学作物生理生态与遗传育种教育部重点实验室,南昌 330045;
    2、江西农业大学农业应对气候变化南昌市重点实验室,南昌 330045
  • 收稿日期:2014-02-24 出版日期:2014-08-18 发布日期:2014-04-16
  • 通讯作者: 黄英金,E-mail:yjhuang_cn@126.com
  • 作者简介:廖江林,E-mail:jlliao514815@163.com
  • 基金资助:

    江西省主要学科带头人项目(7004)、江西省青年科学基金重点项目(20133ACB21004)

Identification of the Differentially Expressed Proteins Between Heat-Tolerant and Heat-Sensitive Rice Responding to High-Temperature Stress at the Early Milky Stage

LIAO  Jiang-Lin-1, 2,SONG  Yu-2, ZHONG  Ping-An-2, ZHOU  Hui-Wen-2, ZHANG  Hong-Yu-2, HUANG  Ying-Jin-1, 2   

  1. 1、Key Laboratory of Crop Physiology, Ecology and Genetic Breeding, Ministry of Education, Jiangxi Agricultural University, Nanchang 330045;
    2、Key Laboratory of Agriculture Responding to Climate Change, Jiangxi Agricultural University, Nanchang 330045
  • Received:2014-02-24 Online:2014-08-18 Published:2014-04-16

摘要: 【目的】鉴定水稻籽粒应答灌浆初期高温胁迫过程中的差异表达蛋白质,并了解其生物学功能。【方法】以耐热水稻纯系XN0437T和热敏感水稻纯系XN0437S为材料,采用桶栽、常规栽培管理方法种植。为保证所取样品生长发育进程一致,在抽穗期标记同一天抽穗的稻穗、在扬花期标记这些稻穗中同一天开花的颖花(稻穗中、下部)。水稻于籽粒灌浆初期(花后第8—14天)移入人工气候箱进行高温(38.0±0.5)℃和常温(25.0±0.5)℃对照处理,处理时间设1、3和5 d;处理结束后,取同一天标记的颖花、采用TCA-丙酮沉淀法提取总蛋白质,用蛋白质双向电泳技术分离获得2个水稻纯系应答灌浆初期高温处理的籽粒蛋白质图谱,采用蛋白质图谱分析软件先分别比较两水稻纯系的处理及其平行对照的蛋白质图谱,确定水稻应答灌浆期高温胁迫中的丰度差异蛋白点,再比较水稻纯系XN0437T和XN0437S应答高温的丰度差异蛋白点,从而确定耐热和热敏感水稻应答高温胁迫的差异表达蛋白点;通过串联质谱分析和数据库搜索鉴定耐热和热敏感水稻应答灌浆初期高温的差异表达蛋白质并分析其涉及的生物学功能。【结果】水稻籽粒蛋白质图谱分析结果显示耐热和热敏感水稻应答灌浆初期高温胁迫过程中存在27个表达丰度相差2倍以上的蛋白点;质谱分析和数据库搜索获得25个差异表达蛋白质,其生物学功能主要涉及生物合成(10个蛋白质)、能量代谢(4个蛋白质)、氧化作用(7个蛋白质)、应激反应(3个蛋白质)和转录调控(1个蛋白质)等生物学过程。【结论】灌浆初期高温胁迫影响水稻籽粒中参与生物合成、能量代谢、氧化作用、应激反应和转录调控等生物学过程相关蛋白质的表达,这些蛋白质的表达模式因水稻基因型(耐热和热敏感水稻)不同、高温胁迫程度(高温处理天数)不同而存在差异。

关键词: 水稻 , 灌浆初期 , 水稻籽粒 , 高温胁迫 , 蛋白质组学

Abstract: 【Objective】The objective of this study is to identify the differentially expressed proteins and their biological functions involving in rice grain responding to high temperature stress at the early milky stage.【Method】The heat-tolerant rice line XN0437T and the heat-sensitive rice line XN0437S were used as plant materials. The tub-planting method and conventional cultivation way were used to culture the rice. To ensure that only uniformly developed samples were used for analysis, rice ears were labeled on the same heading date and the panicles on the labeled ears were further labeled on the same flowering date. On the 10th day after heading, plants with the same label were transferred to chambers and maintained at a temperature of 38.0/25.0 ± 0.5℃ (treatment / control) for 1 day, 3days and 5days. After the initiation of high temperature stress at the early milky stage of rice, rice grains from the same region (middle to bottom part) of labeled ears were harvested at three different time points. The total proteins from harvested sample were extracted by the trichloroacetate (TCA) - acetone precipitation method and the protein profiles of rice grain were set up by two-dimensional electrophoresis (2-DE). The proteins in rice responding to high temperature stress (PRHT) were firstly identified by comparing the 2-DE protein profiles between treatment and its parallel control for heat-tolerant and -sensitive lines, respectively. The differentially expressed proteins were then confirmed by comparing the volume of PRHT from heat-tolerant and -sensitive lines. The functions of these differentially expressed proteins were identified by matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry (MALDI-TOF/TOF MS) analysis and protein database searching. 【Result】Twenty-seven differentially expressed protein spots were found and showed more than 2-folds difference in PRHT volume between the heat-tolerant and -sensitive lines, and 25 proteins were identified by MS and database searching. The 25 identified proteins were involved in biological synthesis (10 proteins), energy metabolism (4 proteins), oxidation (7 proteins), emergency response (3 proteins) and transcriptional regulation (1 protein). 【Conclusion】The expression patterns of the rice grain proteins, which are involved in biological synthesis, energy metabolism, oxidation, emergency response and transcriptional regulation, were influnced by high temperature stress at early milky stage of rice grain. The expression patterns of these proteins were showed variable according to the difference in rice genetype (heat-tolerant and -sensitive lines) and extent (days) of high temperature treatment.

Key words: rice , early milky stage , rice grain , high temperature stress , proteomics