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作物杂志,2024, 第2期: 23–29 doi: 10.16035/j.issn.1001-7283.2024.02.004

• 遗传育种·种质资源·生物技术 • 上一篇    下一篇

苦荞FtERF基因克隆、生物信息学及其表达分析

张俊1(), 蔡苏云1, 徐子豪1, 侯蕾1, 贺润丽1(), 尹桂芳2, 王莉花2(), 王艳青2, 卢文洁2, 孙道旺2   

  1. 1山西中医药大学中药与食品工程学院，030619，山西太原
    2云南省农业科学院生物技术与种质资源研究所/云南省农业生物技术重点实验室/农业农村部西南作物基因资源与种质创制重点实验室，650201，云南昆明
  • 收稿日期:2022-12-30 修回日期:2023-04-26 出版日期:2024-04-15 发布日期:2024-04-15
  • 通讯作者: 贺润丽，研究方向为中药资源开发与利用，E-mail：herunli666@163.com；王莉花，研究方向为植物遗传育种，E-mail：wanglihua70@hotmail.com
  • 作者简介:张俊，研究方向为中药资源开发与利用，E-mail：2639854942@qq.com
  • 基金资助:
    国家自然科学基金(31460379);财政部和农业农村部国家现代农业产业技术体系(CARS-07-C-2);山西省重点研发计划(201803D221012-6);山西省自然科学研究面上项目(20210302123231)

Cloning, Bioinformatics and Expression Analysis of FtERF Gene in Fagopyrum tataricum

Zhang Jun1(), Cai Suyun1, Xu Zihao1, Hou Lei1, He Runli1(), Yin Guifang2, Wang Lihua2(), Wang Yanqing2, Lu Wenjie2, Sun Daowang2   

  1. 1College of Traditional Chinese Medicine and Food Engineering, Shanxi University of Traditional Chinese Medicine, Taiyuan 030619, Shanxi, China
    2Biotechnology and Genetic Germplasm Resources Research Institute, Yunnan Academy of Agricultural Sciences / Yunnan Provincial Key Laboratory of Agricultural Biotechnology / Key Laboratory of Southwestern Crop Gene Resources and Germplasm Innovation, Ministry of Agriculture and Rural Affairs, Kunming 650201, Yunnan, China
  • Received:2022-12-30 Revised:2023-04-26 Online:2024-04-15 Published:2024-04-15

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摘要：

乙烯响应因子对植物发育和细胞代谢具有重要作用。通过RT-PCR从苦荞中克隆出ERF基因，对其进行生物信息学分析和差异表达分析。结果表明，FtERF基因开放阅读框长度为729 bp，编码了243个氨基酸，编码的蛋白分子量26.08 kD，等电点9.22，属于亲水性蛋白。FtERF不含有信号肽和跨膜螺旋结构，亚细胞定位在细胞核内，蛋白质二级结构和三级结构高度相似。FtERF含有抑制基序DLNxxP，系统进化表明FtERF和ERF4关系较近。经荧光定量表达发现，厚壳苦荞不同部位表达均高于薄壳苦荞，苦荞发育成熟期表达高于非成熟期。

关键词: 苦荞, 乙烯响应因子, 生物信息学, 表达分析

Abstract:

Ethylene response factor plays an important role in plant development and cell metabolism. ERF gene was cloned from Tartary buckwheat by RT-PCR, and its bioinformatics and differential expression were analyzed. The results showed that the length of open reading frame of FtERF gene was 729 bp, encoding 243 amino acids, molecular weight of 26.08 kD, isoelectric point of 9.22, belonging to the hydrophilic protein. FtERF didn't contain signal peptides and transmembrane helical structure, and was located in the nucleus of the cell. The secondary and tertiary structures of the proteins were highly similar. FtERF contained the inhibitory motif DLNxxP, and phylogenetic evolution suggested that FtERF and ERF4 were closely related. The results showed that the expression of Tartary buckwheat in different parts of thick-shell were higher than those in thin-shell Tartary buckwheat, and the expression of Tartary buckwheat in mature period was higher than that in immature period.

Key words: Tartary buckwheat, Ethylene response factor, Bioinformatics, Expression analysis

表1

试验所用引物序列信息

引物名称
Primer name		 引物序列
Primer sequence		 用途
Usage
FtERF-F ATGGCACCCAAGCATAAGG 基因克隆
FtERF-R GGCGAGATCTGAAGGCGG
FtERF-F GGTTTCACGGCAGGGATT 实时荧光定量PCR
FtERF-R CACGATAGACGATGAGGCAGA
H3-F AAGAAGTCCCACAGATACCGC
H3-R AGCCTCCTGAAGAGCTAGCAC

图1

FtERF基因PCR产物的电泳结果 M：SM0331 DNA分子量标准；1：云荞1号ERF基因片段；2：小米荞ERF基因片段。

图2

FtERF的保守结构域分析

图3

FtERF的疏水性分析

图4

FtERF的跨膜结构域分析

图5

FtERF的肽信号预测

图6

FtERF的磷酸化位点预测

图7

FtERF蛋白质二级结构预测

图8

FtERF蛋白质三级结构预测

图9

FtERF蛋白的多序列比对 蓝框内为EAR结构域抑制基序。

图10

FtERF与其他物种ERF蛋白的系统进化树

图11

FtERF基因在苦荞不同部位和不同发育时期的相对表达水平 “*”和“**”分别表示在0.05和0.01水平差异显著。

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