《植物生理学报》 2011, 47(5): 488-494

赤桉抗寒转录因子ICE1基因的分子克隆与表达分析

林元震^1,2,3,*, 郭海⁴, 刘纯鑫¹, 陈晓阳¹

1华南农业大学林学院, 广州510642; 2北京林业大学林木花卉遗传育种教育部重点实验室, 北京100083; 3北京林业大学林木 育种国家工程实验室, 北京100083; 4水利部水土保持植物中心, 北京100038

通信作者：林元震；E-mail: yzhlin@scau.edu.cn；Tel: 020-85280259

摘　要：

ICE1属于一种类似MYC的bHLH转录因子, 可特异地结合到CBF3启动子的MYC作用元件并诱导CBF/DREB1下游基 因的转录表达。本文以拟南芥ICE1蛋白序列为信息探针, 搜索桉树基因组和EST数据库的同源序列并进行拼接、设计引 物, 通过RT-PCR从赤桉克隆了桉树的第一个ICE1基因。其cDNA长1 792 bp, 含有完整的开放阅读框, 可编码523个氨基 酸。BLAST分析表明, cDNA序列及其推导的氨基酸序列均与拟南芥、芥菜、小麦和甜杨ICE1存在着较高的同源性, 预示 所获得的cDNA可能是赤桉ICE1基因(EcaICE1)。EcaICE1基因表达分析结果显示, EcaICE1在赤桉根、茎、叶中均表达, 而且表达水平不受低温胁迫处理时间的影响, 这表明EcaICE1是组成型表达。此外, EcaICE1的超表达可以提高转基因烟 草的耐低温能力。上述结果为进一步研究EcaICE1在赤桉耐低温胁迫过程中基因表达的调控机制打下基础。

关键词：赤桉; 转录因子; 抗寒性; ICE1

收稿：2011-01-26   修定：2011-01-26

资助：广东省教育厅育苗工程(LYM10040)、北京林业大学林木育种国家工程实验室开放课题(FOP2010-4)、华南农业大学校长基金(2007K033和2009K014)

Molecular Cloning and Expression Analysis of Cold-Resistant Transcription Factor ICE1 from Eucalyptus camaldulensis L.

LIN Yuan-Zhen^1,2,3,*, GUO Hai⁴, LIU Chun-Xin¹, CHEN Xiao-Yang¹

¹College of Forestry, South China Agricultural University, Guangzhou 510642, China; ²Key Laboratory for Genetics and Breeding in Forest Trees and Ornamental Plants, Ministry of Education, Beijing Forestry University, Beijing 100083, China; ³National Engineering Laboratory for Tree Breeding, Beijing Forestry University, Beijing 100083, China; ⁴Plant Materials for Soil and Water Conservation, Ministry of Water Resources, Beijing 100038, China

Corresponding author: LIN Yuan-Zhen; E-mail: yzhlin@scau.edu.cn; Tel: 020-85280259

Abstract:

ICE1 (inducer of CBF expression 1) is a MYC-like bHLH transcriptional activator, can bind to CBF3 promoter, and induces the expression of CBF3 and its downstream genes during cold acclimation. The first Eucalyptus ICE1 gene was cloned from E. camaldulensis by RT-PCR with the gene-specific primers designed according to Eucalyptus EST and E. grandis genomic sequences and by using Arabidopsis ICE1 protein sequences as the starting information. A 1 792-bp cDNA clone was obtained in our study, which contains a putatively entire open reading frame (ORF) and encodes a MYC-like protein of 523 amino acids. Bioinformatic analysis revealed that this cDNA was highly homologous to ICE1 from Arabidopsis thaliana, Capsella bursa-pastoris, Triticum aestivum and Populus suaveolens, indicating that the interest cDNA was EcaICE1 gene. EcaICE1 was expressed in all tested tissues and the expression levels of EcaICE1 were stable during different low temperature treatments, suggesting that EcaICE1 was expressed constitutively in E. camaldulensis. Moreover, overexpression of EcaICE1 could enhance the cold tolerance in transgenic tobaccos. These results may provide the basic for further study of regulation function of EcaICE1 in gene expression during low temperature stress.

Key words: Eucalyptus camaldulensis; transcription factor; cold resistance; ICE1