小桐子非特异性脂质转移蛋白A的基因克隆及其表达和功能分析

Molecular Cloning, Expression Analysis and Functional Characterization of the Gene Encoding Non-Specific Lipid Transfer Protein A from Jatropha curcas

基于我们最近获得的小桐子低温驯化转录组和数字基因表达谱数据,本工作研究了低温驯化条件下差异表达变化较大的非特异性脂质转移蛋白A基因Jcns LTPA。克隆到该基因的c DNA序列全长833 bp,开放阅读框长度513 bp,编码170个氨基酸,存在ATT_LTSS典型保守功能基序。其启动子区域中鉴定到了TATA框、CAAT框、CATA框、W框等顺式作用元件以及CRT/DRE低温响应元件。半定量RT-PCR分析表明,该基因在茎、根、叶中都有表达,以茎中表达量最高、且受低温诱导最显著。同时,酵母表达Jcns LTPA也提高了重组酵母菌的抗低温能力。这些结果充分说明了小桐子Jcns LTPA是与抗冷性密切相关的基因,可以用于小桐子的抗冷性遗传改良。

From our previous data of the transcriptome and digital gene expression profi ling of cold-acclimated Jatropha curcas, a gene encoding the non-specifi c lipid transfer protein A（Jcns LTPA）, with highly differential expression under chilling, was selectively studied herein. The full length of our cloned Jcns LTPA c DNA is 833 bp, with the entire open reading frame（ORF） of 513 bp encoding a polypeptide of 170 amino acids（with typical ATT_LTSS motif）. Its promoter region was characterized by prediction with TATA-box, CAAT-box, CATA-box, W-box, and the cold responsive elements of CRT/DRE. Semi quantitative RT-PCR analysis revealed that Jcns LTPA was expressed in stem, root, and leaves, with abundant and remarkably chilling-induced expression particularly in stem. Meanwhile, the yeast expression of Jcns LTPA endued the recombinant yeast strain with an enhanced resistance to low temperature. Conclusively, these results largely implicated the cold relevance of Jcns LTPA gene that should be applicable for the genetic improvement on the cold tolerance of J. curcas.