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目的:克隆莲藕CBF2基因的c DNA全长并对其进行序列分析。方法:根据已有的ESTs序列,设计3’和5’端RACE引物,运用RACE技术克隆莲藕CBF2基因c DNA全长。结果:克隆得到全长为1 560bp c DNA序列,其中包括350bp的3’非编码区和124bp的5’非编码区及一个1 086 bp的完整开放阅读框,编码362个氨基酸,序列末端有poly(A)尾。其核苷酸序列与NCBI数据库中大豆DREB2C/CBF2、马铃薯DREB2A/CBF2、黄瓜DREB2A/CBF2及花生DREB2A/CBF2的同源性较高,分别为83%、77%、76%和76%,因此把该基因命名为Lr CBF2。氨基酸序列进化树分析表明该基因与葡萄相关基因亲缘关系较近。结论:分离克隆得到莲藕CBF2基因,为进一步研究莲藕中该基因的功能奠定基础。
OBJECTIVE: To clone the full length cDNA of CBF2 gene of lotus root and analyze its sequence. Methods: According to the existing ESTs sequences, 3 ’and 5’ RACE primers were designed and the full length cDNA of CBF2 gene was cloned by RACE. Results: The full-length cDNA sequence of 1 560 bp was cloned, including 350 bp 3 ’non-coding region, 124 bp 5’ non-coding region and a complete open reading frame of 1 086 bp encoding 362 amino acids with poly (A) tail. The nucleotide sequences showed high homology to DREB2C / CBF2, DREB2A / CBF2, Cucumber DREB2A / CBF2 and DREB2A / CBF2 in NCBI database, which were 83%, 77%, 76% and 76% The gene was named Lr CBF2. Phylogenetic tree analysis of the amino acid sequence showed that the gene was closely related to grape-related genes. Conclusion: The CBF2 gene of lotus root is isolated and cloned, which lays the foundation for further study on the function of this gene in lotus root.