遗传 ›› 2020, Vol. 42 ›› Issue (8): 788-798.doi: 10.16288/j.yczz.20-141

• 研究报告 • 上一篇    下一篇

大豆E2泛素结合酶基因GmUBC1的克隆及在拟南芥中的异源表达

毛卓卓, 宫宇, 史贵霞, 李亚丽, 喻德跃, 黄方   

  1. 南京农业大学农学院,作物遗传与种质创新国家重点实验室,国家大豆改良中心,农业部大豆生物学与遗传育种实验室(综合性),南京 210095
  • 收稿日期:2020-05-21 修回日期:2020-07-07 出版日期:2020-08-20 发布日期:2020-08-07
  • 作者简介:毛卓卓,在读硕士研究生,专业方向:作物遗传育种。
  • 基金资助:
    国家自然科学基金项目资助编号(31871649)

Cloning of the soybean E2 ubiquitin-conjugating enzyme GmUBC1 and its expression in Arabidopsis thaliana

Mao Zhuozhuo, Gong Yu, Shi Guixia, Li Yali, Yu Deyue, Huang Fang   

  1. State Key Laboratory of Crop Genetics & Germplasm Enhancement, National Center for Soybean Improvement, Key Laboratory of Biology and Genetic Improvement of Soybean, Ministry of Agriculture, Nanjing Agricultural University, Nanjing 210095, China
  • Received:2020-05-21 Revised:2020-07-07 Online:2020-08-20 Published:2020-08-07
  • Supported by:
    Supported by the National Natural Science Foundation of China No(31871649)

摘要:

E2泛素结合酶(ubiquitin-conjugating enzyme)参与植物的抗逆、生长发育等多种生物途径的调控,其功能研究在拟南芥(Arabidopsis thaliana)中的报道较多,但在重要经济作物大豆(Glycine max)中鲜有报道。本研究从大豆“南农94-16”中克隆了1个与大豆子叶折叠突变体可能相关的基因Glyma.12G161200,序列分析结果表明该基因编码一个E2泛素结合酶,因此将其命名为GmUBC1。该基因编码区全长为462 bp,编码一个含153个氨基酸的蛋白,预测其分子量为17.25 kDa,等电点为6.74。利用qRT-PCR技术对GmUBC1在大豆不同组织中的表达模式及其对不同非生物胁迫和激素处理的响应模式进行分析,发现该基因在开花后40 d种子中表达量最高,在PEG、低温、JA和ABA处理下表达下调。亚细胞定位分析发现GmUBC1蛋白在整个细胞内都有表达。进一步在拟南芥中异源表达GmUBC1,发现转基因株系的千粒重和总氨基酸含量显著提高,表明异源过表达GmUBC1可以调控种子重量和氨基酸含量,为大豆品质改良提供了基因资源。

关键词: 大豆, E2泛素结合酶, GmUBC1, 异源表达

Abstract:

Plant E2 Ubiquitin-conjugating enzymes regulate various biological pathways such as stress resistance, growth and development. Reports on its functions are more frequent in Arabidopsis thaliana, but relatively rare in soybean (Glycine max), which is one of the most important economic crops. In this study, a gene Glyma.12G161200, which may be related to the soybean cotyledon folding mutant, was cloned from soybean “Nanong 94-16”. Analysis of its sequence suggested that it encodes an E2 ubiquitin binding enzyme, so it was named as GmUBC1. Its coding region is 462 bp in length, which encodes a protein of 153 amino acids with a predicted molecular mass of 17.25 kDa and an isoelectric point of 6.74. The expression pattern of GmUBC1 in different tissues of soybean and its response patterns to different stresses and hormone treatments were analyzed by real-time PCR. The results showed that the gene was expressed at the highest level in mutant seeds at 40 days after flowering. Moreover, the expression of the GmUBC1 gene was down-regulated by the treatments of PEG, cold, JA and ABA, respectively. Subcellular localization analysis of GmUBC1 revealed that the protein was expressed in the whole cell. When GmUBC1 was ectopically expressed in Arabidopsis, the 1000-grain weight and total amino acid content of some transgenic lines were found to be significantly increased. Collectively, heterologous overexpression of GmUBC1 can regulate seed weights and amino acid contents, which may provide genetic resources for soybean quality improvement.

Key words: soybean (Glycine max L.), ubiquitin-conjugating enzyme E2, GmUBC1, heterologous expression