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作物学报 ›› 2019, Vol. 45 ›› Issue (5): 693-704.doi: 10.3724/SP.J.1006.2019.84120

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

人工合成甘蓝型油菜抗旱性及DNA甲基化水平分析

袁溢,朱双,方婷婷,蒋金金(),王幼平   

  1. 扬州大学生物科学与技术学院, 江苏扬州 225009
  • 收稿日期:2018-09-09 接受日期:2018-12-24 出版日期:2019-05-12 网络出版日期:2019-01-17
  • 通讯作者: 蒋金金
  • 基金资助:
    本研究由国家自然科学基金项目(31771824);本研究由国家自然科学基金项目(31771825);中国和江苏省博士后科学基金项目(2014M561719);中国和江苏省博士后科学基金项目(2015T80591);中国和江苏省博士后科学基金项目(1401078B)

Analysis of drought resistance and DNA methylation level of resynthesized Brassica napus

Yi YUAN,Shuang ZHU,Ting-Ting FANG,Jin-Jin JIANG(),You-Ping WANG   

  1. College of Bioscience and Biotechnology, Yangzhou University, Yangzhou 225009, Jiangsu, China
  • Received:2018-09-09 Accepted:2018-12-24 Published:2019-05-12 Published online:2019-01-17
  • Contact: Jin-Jin JIANG
  • Supported by:
    This study was supported by the National Natural Science Foundation of China(31771824);This study was supported by the National Natural Science Foundation of China(31771825);China and Jiangsu Postdoctoral Science Foundation(2014M561719);China and Jiangsu Postdoctoral Science Foundation(2015T80591);China and Jiangsu Postdoctoral Science Foundation(1401078B)

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摘要：

甘蓝型油菜是具有重要经济价值的多倍体物种, 是优质食用植物油和饲料蛋白质的重要来源之一。但是其驯化历史较短, 遗传背景狭窄, 且在整个生命周期中都对干旱胁迫敏感, 因此培育高产耐旱品种是甘蓝型油菜的重要育种目标之一。本文用15% PEG-6000模拟干旱胁迫, 对人工合成甘蓝型油菜不同世代(S1~S4)及其二倍体亲本进行不同时间的胁迫处理, 并结合表型观察, 以及叶片中丙二醛(MDA)、可溶性蛋白含量、过氧化物酶(POD)、超氧化物歧化酶(SOD)等生理指标的测定, 初步了解上述材料的抗旱性差异。结合表型观察和叶片中相对含水量分析, 发现人工合成甘蓝型油菜S1~S4及其亲本的抗旱性表现为甘蓝 > Bn-S3 > Bn-S4 > Bn-S1 > Bn-S2 > 白菜型油菜。干旱胁迫后Bn-S3、Bn-S4的POD及SOD活性较高, MDA含量较低, 表明Bn-S3和Bn-S4能更加有效地清除活性氧(ROS), 对过氧化损伤的防御能力更强。通过HPLC分析发现所有材料的甲基化水平在胁迫12 h时最高, 其中亲本白菜型油菜Br的甲基化水平最高, Bn-S1和Bn-S4介于两亲本之间, 而Bn-S2和Bn-S3低于两亲本。甲基化敏感多态性分析也显示人工合成甘蓝型油菜在干旱胁迫后, 甲基化和去甲基化水平均发生了明显的变化, 表明植物的甲基化变化可能有利于提高其抗旱能力。

关键词: 人工合成甘蓝型油菜, 白菜型油菜, 甘蓝, 干旱胁迫, DNA甲基化

Abstract:

Brassica napus, as one of the important resources of edible plant oil and forage protein, is a polyploid species with great economic value. However, it is sensitive to drought stress throughout whole lifecycle due to short domestication history and narrow genetic background. Thus, it is a main purpose to breed B. napus cultivar with high yield and drought resistance. In the present study, we compared the drought resistance among S1-S4 generations of resynthesized B. napus and diploid parents under different time periods of 15% PEG-6000 treatment. The different drought tolerance levels were assessed based on phenotype observation, leaf physiological indexes (MDA, soluble protein, SOD and POD). Accompanying with water content analysis, we found the drought tolerance showed a trend of B. oleracea > Bn-S3 > Bn-S4 > Bn-S1 > Bn-S2 > B. rapa. Under drought stress, POD and SOD activities in Bn-S3 and Bn-S4 were higher than these in other plants tested, and MDA content was decreased, indicating that Bn-S3 and Bn-S4 have better ability in clearing ROS, and defending from peroxidation damage. On the basis of HPLC analysis, the methylation level in all materials was the highest under drought stress of 12 h. And the methylation level in B. rapa was higher than that in others, that in Bn-S1 and Bn-S4 was between that in parents, while that in Bn-S2 and Bn-S3 was lower than that in parents. Methylation sensitive amplification polymorphism analysis also revealed multiple changes in methylation and demethylation level of resynthesized B. napus under drought stress, indicating methylation changes might be involved in plant drought tolerance.

Key words: resynthesized Brassica napus, B. rapa, B. oleracea, drought stress, DNA methylation

图1

干旱胁迫后植物的表型 Br: 白菜型油菜; Bo: 甘蓝; S1~S4: 人工合成甘蓝型油菜自交后代。"

图2

干旱胁迫12 h后植物的相对含水量和相对保水力 CK: 对照; DT: 干旱胁迫; Br: 白菜型油菜; Bo: 甘蓝; S1~S4: 人工合成甘蓝型油菜自交后代。"

图3

干旱胁迫处理下植物的生理指标分析 A: 过氧化物酶活力; B: 丙二醛含量; C: 超氧化物歧化酶活力; D: 可溶性蛋白含量。平均值±标准误, n = 3。Br: 白菜型油菜; Bo: 甘蓝; S1~S4: 人工合成甘蓝型油菜自交后代。"

图4

干旱胁迫处理下植物DNA甲基化水平的变化 Br: 白菜型油菜; Bo: 甘蓝; Bn-S1~S4: 人工合成甘蓝型油菜自交后代。"

图5

干旱胁迫处理下植物中不同类型DNA甲基化修饰的百分比 Br: 白菜型油菜; Bo: 父本甘蓝; Bn-S1~S4: 人工合成甘蓝型油菜自交后代。"

表1

干旱胁迫6 H植物甲基化水平的变化"

表2

干旱胁迫12H植物甲基化水平的变化"

表3

干旱胁迫24H植物甲基化水平的变化"

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