花生中DREB类转录因子PNDREB1的克隆及鉴定  

Isolation and Identification of PNDREB1:A New DREB Transcription Factor from Peanut(Arachis hypogaea L.)

作　　者：张梅[1,2,3] 刘炜[1,2] 毕玉平[1,2,3] 王自章[4] 

机构地区：[1]山东省农业科学院高新技术研究中心/山东省作物与畜禽品种改良生物技术重点实验室,山东济南250100 [2]农业部黄淮海作物遗传改良与生物技术重点开放实验室,山东济南250100 [3]山东师范大学生命科学学院,山东济南250014 [4]中国科学院植物研究所,北京100093

出　　处：《作物学报》2009年第11期1973-1980,共8页Acta Agronomica Sinica

基　　金：转基因生物新品种培育科技重大专项(2009ZX08001-010B);山东省农业科学院博士基金项目(2007YBS008)资助

摘　　要：脱水响应元件结合因子(DREB)是一类对多个抗逆相关基因表达起调控作用的植物特有的转录因子,在植物抗逆能力综合改良方面具有重要作用。本研究通过筛选花生未成熟种子的cDNA文库,分离到一个DREB类基因——PNDREB1(FM955398)。该基因序列长度为687bp,推测编码蛋白含有229个氨基酸残基,相对分子量为24.7kD,理论等电点为5.97,与其他物种中该类转录因子序列同源性较高。利用酵母单杂交系统,对花生PNDREB1与DRE元件的特异识别和结合能力及其C-末端转录激活活性进行检测,显示,转入含有该基因及阳性对照基因的载体均可使酵母菌株正常生长,且具有显色反应,而转入空载体的酵母菌株不能生长,证明基因PNDREB1中的AP2结构域具有和DRE元件特异性结合的能力;同时,只有转入含有该基因C-末端片段及阳性对照基因的载体可使酵母菌株正常生长,并具有显色反应,而转入空载体的酵母菌株不能生长,说明其C-末端片段具有转录激活活性,该基因为DREB类转录因子。基因表达模式分析显示,PNDREB1为组成型表达,且被低温强烈、迅速诱导表达,并对干旱胁迫也有一定程度的响应,但对高盐和ABA处理没有响应。The dehydration responsive element binding proteins （DREB） are important and specific plant transcription factors responding to stress conditions including drought, salt and low temperature. It has been generally accepted that DREB can regulates the expression of a number of abiotic stress-related genes in down stream of the stress signal transduction pathways. In this paper, a DREB-like gene, named PNDREB1 （accession No. FM955398）, was cloned by screening a peanut （Arachis hypogaea L.） full-length cDNA library of immature seeds. The structure analysis showed that PNDREB1 contained a 687 bp ORF, encoding a protein of 229 amino acids with predicted molecular weight of 24.7 kD and a isoelectric point of 5.97. The predicted protein sequence contained one conserved AP2 domain, which is the typical characteristic of DREB transcription factors. Based on the sequences similarity, PNDREB1 is classified into A-1 subgroup of DREB subfamily. Furthermore, the yeast hybrid system was carried out, and the results confirmed that the AP2 domain of PNDREB1 could specifically interact with DRE cis-acting element. The activation activity of the C-terminal end as a transcriptional activator was also been proved experimentally. The expression pattern analysis carried out by semi-quantitative RT-PCR indicated that PNDREB1 was constitutively expressed in various tissues of peanut, and was strongly upregulate by treatments with low temperature, also respond to dehydration. However, the expression of PNDREB1 was not affected by high salinity and exogenous application of abscisic acid （ABA）. In this study, we isolated and characterized a novel peanut DREB-like transcription factor which was regulated by low temperature and osmotic stresses.

关 键 词：花生 DREB类转录因子 PNDREB1 酵母单杂交 表达模式 低温 干旱 

分 类 号：S565.2[农业科学—作物学]