山茱萸CoDXR基因的克隆与分析

Cloning and analysis of CoDXR gene in Cornus officinalis

目的克隆山茱萸关键酶基因1-脱氧-D-核酮糖5-磷酸还原酶(CoDXR)的全长cDNA序列,为山茱萸的进一步研究提供依据。方法以本实验室获得的山茱萸转录组数据中的转录本序列c147202_g1为模板,通过Primer Premier 5.0设计特异性引物,利用RT-PCR技术克隆CoDXR基因的全长cDNA序列,并通过相关的生物信息学软件对其进行相关的生物信息学分析和功能预测。结果 CoDXR基因长度为1 505 bp,ORF长度是729 bp,编码242个氨基酸。而通过SignalP4.0Server和HMMTOP对CoDXR蛋白预测分析结果可知,该蛋白不具有信号肽和跨膜区,为疏水性蛋白。系统进化树分析结果表明CoDXR蛋白与野茶树(Camelliasinensis)的DXR蛋白序列相似性最高。结论在山茱萸果实和叶片转录组测序的基础上成功克隆了山茱萸萜类合成途径中的关键酶基因CoDXR,对其进行相关的生物信息学分析,为深入研究CoDXR基因在山茱萸萜类合成途径中的功能奠定了基础。

Objective To clone the full-length cDNA sequence of CoDXR, a key enzyme gene of Cornus officinalis, and provide a basis for further study of C. officinalis. Methods In this study, we used the transcript sequence c147202_g1 from the transcriptome data of C. officinalis obtained in our laboratory as template, designed specific primers through Primer Premier 5.0, cloned the full-length cDNA sequence of C. officinalis DXR gene by RT-PCR technology, and the bioinformatics analysis and function prediction were carried out through the relevant bioinformatics software. Results The results showed that the CoDXR gene was 1 505 bp in length and the ORF was 729 bp in length, encoding 242 amino acids. The results of predictive analysis of CoDXR protein by SignalP4.0 Server and HMMTOP showed that the protein was a hydrophobic protein without signal peptide and transmembrane region. Phylogenetic tree analysis showed that the CoDXR protein had the highest similarity to the DXR protein sequence of Camellia sinensis. Conclusion In this study, the key enzyme gene CoDXR was successfully cloned based on the sequencing of the C. officinalis transcriptome, and related bioinformatics analysis was carried out. The results of this study laid the foundation for further study on the function of CoDXR gene in the terpenoid synthesis pathway of C. officinalis.