茶树CsCBF1基因克隆和转录活性分析

Isolation and Transcription Activation Analysis of the CsCBF1 Gene fromCamellia sinensis

采用RACE技术克隆了一个受冷诱导的茶树CBF基因全长cDNA,命名为CsCBF1(GenBank登录号为EU563238)。CsCBF1cDNA全长序列为1 211bp,开放阅读框编码259个氨基酸。氨基酸序列分析表明,CsCBF1具有CBF家族典型的保守结构域,与其他植物的CBF具有较高的相似性;与拟南芥、辣椒和橡胶树编码的CBF相似性分别为56%、63%和56%。亚细胞定位结果表明,CsCBF1位于细胞核内。分别将10个CsCBF1缺失突变体与GAL4DNA结合域融合的结果显示,CsCBF1的羧基末端酸性结构域(第137位氨基酸至259位氨基酸)在酵母中具有转录激活活性。实时定量RT-PCR分析表明,CsCBF1基因受低温的快速诱导表达。

The full-length cDNA of a cold-induced CBFgene was cloned from tea（Camellia sinensis）,and designated as CsCBF1（GenBank accession No.EU563238）by RACE method.The cDNA was 1 211 bp in length,and contained an open reading frame encoding 259 amino acids.Amino acid sequence alignment showed that CsCBF1 contained all typical conserved motifs of CBF family.CsCBF1 protein was very similar in amino acid sequence to CBFs from other plant species,and showing 56%,63% and 56% amino acid sequence identity to CBFs fromArabidopsis thaliana,Capsicum annuum,and Hevea brasiliensis,respectively.Subcellular location assay indicated that CsCBF1-GFP fusion protein was located in nucleus.When ten deletion mutants of CsCBF1 were fused in-frame to the yeast GAL4DNA-binding domain,its carboxyl-terminal acidic region（from 137th amino acid to 259th amino acid）could activate the transcription in yeast.Quantitative real-time RT-PCR analysis revealed that CsCBF1 was induced quickly by low temperature.