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作物学报 ›› 2021, Vol. 47 ›› Issue (10): 2045-2052.doi: 10.3724/SP.J.1006.2021.02079

• 研究简报 • 上一篇    下一篇

水稻基因OsATS的克隆及功能鉴定

李晓旭(), 王蕊(), 张利霞, 宋亚萌, 田晓楠, 葛荣朝*()   

  1. 河北师范大学生命科学学院, 河北石家庄 050024
  • 收稿日期:2020-11-21 接受日期:2021-03-22 出版日期:2021-10-12 网络出版日期:2021-04-07
  • 通讯作者: 葛荣朝
  • 作者简介:李晓旭, E-mail: jiandanxiaoxiao@163.com;
    王蕊, E-mail: 1915435558@qq.com.
    第一联系人：

    ** 同等贡献(Contributed equally to this work)

  • 基金资助:
    国家自然科学基金项目(30900104);河北省自然科学基金项目(C2016205158)

Cloning and functional identification of gene OsATS in rice

LI Xiao-Xu(), WANG Rui(), ZHANG Li-Xia, SONG Ya-Meng, TIAN Xiao-Nan, GE Rong-Chao*()   

  1. College of Life Science, Hebei Normal University, Shijiazhuang 050024, Hebei, China
  • Received:2020-11-21 Accepted:2021-03-22 Published:2021-10-12 Published online:2021-04-07
  • Contact: GE Rong-Chao
  • Supported by:
    National Natural Science Foundation of China(30900104);Natural Science Foundation of Hebei Province(C2016205158)

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摘要：

植物胚胎特异性蛋白ATS3和植物的渗透胁迫响应有密切关系, 本文对水稻OsATS基因的抗逆相关功能进行了初步研究。qRT-PCR检测发现, 水稻在盐胁迫后OsATS基因表达量显著增加。构建OsATS基因过表达载体, 转化拟南芥植株, 抗逆性检测表明, OsATS基因的过表达可以显著提高拟南芥在萌发阶段和成株阶段的耐盐性。随后将过表达载体p1300-35S:OsATS和RNA干涉载体pTCK303-OsATS-RNAi转入水稻, 抗逆性分析表明, OsATS过表达水稻株系在萌发阶段和苗期的耐盐性显著提高, 而OsATS基因RNAi水稻株系耐盐性则明显下降。qRT-PCR和生理指标检测表明, OsATS基因的表达可能通过调节OsP5CS1、OsLEA3-1、OsPDH基因的表达, 调控了水稻细胞中的脯氨酸、LEA蛋白质含量, 进而影响了水稻植株整体的耐盐性。本研究初步揭示了OsATS基因的抗逆功能, 后续可通过调整该基因的表达量, 改良水稻的抗逆性。

关键词: 水稻, OsATS基因, 过表达, RNAi干扰, 生理指标

Abstract:

The plant embryo specific protein ATS3 is closely related to osmotic stress response in plants. Here, the stress resistance related gene OsATS was preliminarily studied in rice. Fluorescence quantitative PCR showed that the relative expression level of OsATS increased significantly after salt stress in rice. The overexpression vector of OsATS was constructed and transformed into Arabidopsis thaliana. The stress resistance test revealed that the overexpression of OsATS gene could significantly improve the salt tolerance of Arabidopsis thaliana at germination and adult stages. After that, the overexpression vector p1300-35s:OSATS and RNA interference vector pTCK303-OsATS-RNAi were transferred into rice. The stress tolerance analysis indicated that the salt tolerance of OsATS overexpression rice lines significantly increased at germination stage and seedling stage, while the salt tolerance of OsATS RNAi rice lines significantly decreased. Results of qRT-PCR and physiological index detection demonstrated that the relative expression levels of OSATS gene might regulate the protein content of proline and LEA cells by regulating the expression of OsP5CS1, OsLEA3-1 and OsPDH, thus affecting the salt tolerance in rice. This study preliminarily revealed the stress resistance function of OSATS gene, which laid a foundation for improving rice stress resistance by adjusting the relative expression level of OSATS gene.

Key words: rice, OsATS genes, overexpression, RNA interference, physiological indexes

图1

盐胁迫后水稻OsATS基因表达量的变化 差异显著性分析采用t-test 方法, *P < 0.05、**P < 0.01。"

图2

OsATS在烟草表皮的亚细胞定位"

图3

OsATS过表达拟南芥的PCR鉴定"

图4

OsATS基因表达量的qRT-PCR检测 差异显著性分析采用t-test方法, *** P <0.001。"

图5

盐胁迫下OsATS过表达拟南芥种子萌发率统计 差异显著性分析采用t-test方法, * P < 0.05、** P < 0.01。"

图6

OsATS过表达拟南芥成株的耐盐性检测"

图7

OsATS过表达水稻和RNAi转基因水稻的qRT-PCR检测 WT: 野生型水稻; OX: OsATS过表达转基因水稻株系; Ri: RNAi转基因水稻株系; 差异显著性分析采用t-test方法, *** P < 0.001。"

图8

OsATS过表达水稻和RNAi转基因水稻种子在NaCl胁迫下的萌发率"

图9

OsATS转基因水稻的耐盐性检测"

图10

OsATS过表达和RNAi水稻的生理指标检测 A: 脯氨酸含量; B: 丙二醛含量; C: 细胞质膜透性。差异显著性分析采用t-test方法, * P < 0.05、** P < 0.01、*** P <0.001。"

图11

OsATS转基因水稻中耐盐相关基因的表达变化 差异显著性分析采用t-test方法, * P < 0.05、** P < 0.01、*** P < 0.001。"

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