冬凌草1-脱氧木酮糖-5-磷酸还原异构酶(DXR)基因克隆与分析

Cloning and Analysis of c DNA Encoding Key Enzyme DXR in Diterpenoids Biosynthesis Pathway from Rabdosiae Rubescentis Herba

目的:为研究冬凌草二萜类合成的相关基因,在冬凌草转录组信息数据的基础之上,以冬凌草无菌苗为研究材料,克隆冬凌草二萜类合成的关键酶1-脱氧木酮糖-5-磷酸还原异构酶(l-deoxy-D-lxyluloses-phosphatereduetoisomerase,DXR)基因。方法:采用逆转录PCR技术克隆冬凌草DXR基因,实时荧光定量PCR法分析其组织表达模式。结果:DXR c DNA基因全长1 500 bp,DXR基因开放阅读框为1 422 bp,编码473个氨基酸组成的蛋白质序列,理论相对分子质量为51.39 k Da,等电点为6.09,是一种亲水性蛋白。DXR在茎中表达量相对较高,在愈伤组织中表达量最低。结论:研究结果为深入研究冬凌草DXR酶的活性和功能及为冬凌草二萜类化合物的生物合成机制、优良基因挖掘奠定基础。

Objective： To study the genes related to the synthesis of diterpenoid. Based on the data of transcriptome sequencing, c DNAs encoding 1-deoxy-D-xylulose-5-phosphatereduetoisomerase（ DXR） were obtained from the leaves of aseptic seedlings of Rabdosiae Rubescentis Herba. Method： DXR was obtained by reverse transcription PCR. Real-time quantitative PCR was used to detect the relative expression patterns of DXR in different tissues of Rabdosiae Rubescentis Herba. Result： Sequence analysis showed that the full-length c DNA of DXR was 1 500 bp and contains gene open reading frame（ ORF） of 1 422 bp encoding 473 amino acids. The theoretical molecular weight was 51. 39 k Da and the isoelectric point was predicted as 6. 09,suggesting it was a type of hydrophilic protein. The expression pattern of the gene in different tissues was analyzed by Real-time fluorescence quantitative PCR. The results showed the expression of DXR was relatively high in the stem and the lowest in callus. Conclusion： The results will provide a basis for studying the activity and function of DXR from Rabdosiae Rubescentis Herba,and lay a foundation for biosynthesis and gene mining of terpenoids.