花椰菜BraERF023a的克隆及在响应盐和干旱胁迫中的功能

doi:10.3864/j.issn.0578-1752.2021.01.011

摘要/Abstract

摘要：

【目的】在前期从花椰菜中筛选、鉴定出多个逆境胁迫应答相关ERF（Ethylene-responsive factors）转录因子的基础上,对其中一个未知功能的转录因子BraERF023a进行克隆,并阐释其在盐及干旱胁迫条件下的表达特征及功能。【方法】对花椰菜中的BraERF023a进行克隆,采用MEGA 6等生物学软件对其序列特征进行分析,采用qRT-PCR方法分析BraERF023a在盐及干旱胁迫条件下的表达特征,进而构建过表达载体并转化拟南芥,对BraERF023a过表达转基因拟南芥株系在盐及干旱胁迫下的表型、生存率进行观察、分析。【结果】序列分析证实,花椰菜BraERF023a编码区长度为597 bp,编码含198个氨基酸的蛋白质,其内含有一个AP2保守结构域。BraERF023a在十字花科芸薹属植物中具有很高保守性。转录表达模式分析显示,盐及干旱胁迫条件均可显著诱导花椰菜BraERF023a的表达。进一步的功能分析显示,在盐及干旱胁迫处理条件下,过表达BraERF023a的转基因拟南芥株系比野生型对照生长势更好,植株生存率更高,盐及干旱耐受能力明显增强。【结论】BraERF023a在花椰菜响应盐及干旱胁迫中发挥重要的正向调控作用,过表达BraERF023a可显著提高转化株对盐及干旱的胁迫耐受。

关键词: 花椰菜, ERF转录因子, BraERF023a, 盐胁迫, 干旱胁迫

Abstract:

【Objective】In our previous study, Ethylene-responsive factors (ERFs) involved in abiotic stress responses in cauliflower were identified. In the present study, one of these ERF transcription factors with unknown function, named BraERF023a, was cloned. The expression profiles and function of BraERF023a under salt and drought stresses were further explored. 【Method】BraERF023a was cloned by RT-PCR in cauliflower. The sequence characteristics of BraERF023a were analyzed by biosoftwares such as MEGA 6. The expression profiles of BraERF023a under salt and drought stresses were explored by qRT-PCR. BraERF023a overexpression vector was then constructed and genetically transformed into Arabidopsis. The phenotypes and survival rate of overexpression BraERF023a transgenic Arabidopsis under salt and drought stresses were observed and analyzed.【Result】Sequence analysis confirmed that BraERF023a coding region was 597 bp in length and encoded a protein containing 198 amino acids with an AP2 conserved domain. BraERF023a was highly conserved in Brassica. The expression level of BraERF023a significantly increased under both salt and drought stresses in cauliflower. Functional analysis indicated that comparative with the wild-type controls, overexpression BraERF023a transgenic Arabidopsis lines exhibited better growth vigor and higher plant survival rate under salt or drought stress.【Conclusion】 BraERF023a played important roles in positive response to salt and drought stresses in cauliflower. Overexpression of BraERF023a in Arabidopsis could significantly improve the tolerance of transgenic lines to salt and drought stresses.

Key words: cauliflower, ERF transcription factor, BraERF023a, salt stress, drought stress

李慧,韩占品,贺丽霞,杨亚苓,尤书燕,邓琳,王春国. 花椰菜BraERF023a的克隆及在响应盐和干旱胁迫中的功能[J]. 中国农业科学, 2021, 54(1): 152-163.

LI Hui,HAN ZhanPin,HE LiXia,YANG YaLing,YOU ShuYan,DENG Lin,WANG ChunGuo. Cloning and Functional Analysis of BraERF023a Under Salt and Drought Stresses in Cauliflower (Brassica oleracea L. var. botrytis)[J]. Scientia Agricultura Sinica, 2021, 54(1): 152-163.

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引物名称 Primer name	引物序列 Primer sequence (5′-3′)
BraERF023a-F	ATGCTGAGTACAGTGAGTGAAACC
BraERF023a-R	TCACAGACACGCCATGAACTG
Nco I-BraERF023a-F	CCATGGATGCTGAGTACAGTGAGTG
BstE II-BraERF023a-R	GGTCACCTCACAGACACGCCAT
qBraERF023a-F	CCACCACTTACCAACGAAC
qBraERF023a-R	AAGATCCGAGCCAAATCCG
35S-F	AACAGAACTCGCCGTAAAG
BraActin-F	GCTCCTCTTAACCCAAAGGC
BraActin-R	CACACCATCACCAGAATCCAGC
AtActin-F	GGTAACATTGTGCTCAGTGGTGG
AtActin-R	AACGACCTTAATCTTCATGCTGC

表型Phenotype	对照CK	L4	L8	L9
株高 Stem length (cm)	43.125±2.780c	56.750±3. 862a	51.125±1.0307ab	49.625±3.0923b
抽薹时间 Bolting time (d)	26.00±2.4495b	32.00±2.4495a	29±1.9148a	31.00±1.2583a
茎粗 Stem diameter (mm)	1.2375±0.0727b	1.7800±0.2191a	1.6300±0.1551a	1.7125±0.1059a
主茎 Stem (number)	5.75±0.9574c	6.75±1.0708ab	6.25±0.5000b	7.775±0.5774a
一级分枝 Primary branch (number)	13.75±2.630b	15.00±2.449ab	16.00±3.742a	13.75±5.132b
二级分枝 Secondary branch (number)	5.00±0.816a	5.00±1.155a	5.75±2.602a	6.00±1.15a
果荚长 Pod length (mm)	18.000±1.0475a	17.153±1.3553a	17.600±1.2035a	18.028±0.8587a