中间锦鸡儿CiDR1的克隆及干旱胁迫下的表达分析

Cloning of CiDR1 from Caragana intermedia and Its Expression under Drought Stress

[目的]研究并了解中间锦鸡儿CiDR1基因功能及其对干旱胁迫的响应,为抗性育种提供候选基因。[方法]通过RACE技术从中间锦鸡儿中克隆CiDR1基因的c DNA全长,利用生物信息学分析软件对其基因结构及功能进行分析和预测。再通过qRT-PCR技术对干旱胁迫后的幼苗中的CiDR1表达模式进行研究。[结果]从中间锦鸡儿中克隆到CiDR1基因的c DNA全长共计4 297 bp,Gen Bank登录号为KP277100。生物信息学分析表明,预测的CiDR1蛋白序列中含有1 243个氨基酸残基,具有抗病基因特征结构域TIR、NB-ARC、LRR等,其等电点为6.35,不稳定指数为42.91,不具备信号肽,为非分泌蛋白,定位于细胞质中。定量PCR检测发现,CiDR1基因在幼年期的茎中表达量较低,在成年期的叶片中表达量较高;在干旱胁迫处理后的幼苗中,CiDR1表达水平有明显下降,表明该基因的表达受干旱抑制,可能与中间锦鸡儿适应干旱相关。[结论]中间锦鸡儿在干旱胁迫后其根、茎和叶中CiDR1的表达均明显下降,表明CiDR1的表达受干旱抑制,可能与中间锦鸡儿适应干旱相关,进一步研究发现CiDR1在根、茎、叶中的表达水平可能受发育阶段调控。

To study the function of CiDR1 gene in Caragana intermedia and its response to drought stress.[Method]The full-length cDNA sequence of CiDR1 gene was cloned using RACE methods, and then the bioinformatics analysis were carried out to predict gene structure. Finally, CiDR1 expression patterns under drought stress was investigated by qRT-PCR. [Result]The full-length cDNA sequence of CiDR1 gene was 4 297 bp. The deduced amino-acid sequence for CiDR1 was 1 243 amino acids long and contained toll/interluekin receptor （TIR）, nucleotide binding site （NBS） and leucine-rich repeats （LRR） motif, which are the characteristics of plant resistance genes. The theoretical isoelectric point and instability index of CiDR1 protein was 6.35 and 42.91, respectively. Further analysis showed that CiDR1 protein had no signal peptide, was a non-secretary protein and located in the cytoplasmic matrix. The qPCR analysis showed that the CiDR1 transcripts were expressed strongly in adult leaves and weakly in juvenile stems, and after drought treatments the levels of CiDR1 transcripts decreased. [Conclusion]The results indicate that CiDR1 might play an role in response of C. intermedia to drought and could be used for molecular breeding.