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雷公藤4—（5′—二磷酸胞苷）—2—C—甲基—D—赤藓醇激酶基因的全长克隆与表达分析水晶白坯 4-（5-焦磷酸胞苷）-2-C-甲基-D-赤藓醇激酶[4-（cytidine-5-diphospho）-2-C-methyl-D-erythritol kinase， CMK]是雷公藤甲素等萜类成分生物合成途径上关键酶之一。根据获得的雷公藤转录组数据中TwCMK片段设计特异性引物，采用全长PCR方法克隆TwCMK全长cDNA，并进行生物信息学分析；采用实时荧光定量PCR （RT-qPCR）检测茉莉酸甲酯 （MeJA） 诱导雷公藤悬浮细胞后0～72 h TwCMK基因的表达水平。结果表明，TwCMK全长cDNA由1 732个核苷酸组成，具有完整的开放阅读框，共编码387个氨基酸，蛋白相对分子质量为42.85 kDa，理论等电点为5.79；TwCMK基因受MeJA诱导后表达逐渐上升，在24 h时达到最高值。为进一步研究雷公藤甲素等萜类成分次生代谢调控和生物合成奠定了基础。 胞苷酸标签： 雷公藤；4-（5-焦磷酸胞苷）-2-C-甲基-D-赤藓醇激酶（CMK）；生物信息学分析；表达分析311图钉
[Abstract] 4-（Cytidine-5-diphospho）-2-C-methyl-D-erythritol kinase is a key enzyme in the biosynthesis pathway of terpenoids. According to the transcriptome database， the specific primers were designed and used in PCR. The bioinformatic analysis of the sequenc
ed TwCMK gene was performed in several bioinformatics software.The Real-time fluorescence quantification polymerase chain reaction （RT-qPCR） were used to detect the expression levels of TwCMK from T. wilfordii after elicitor MeJA supplied. The results showed that the full length of TwCMK cDNA was 1 732 bp encoding 387 amino acids. The theoretical isoelectric point of the putative TwCMK protein was 5.79 and the molecular weight was about 42.85 kDa. MeJA stimulated the rising of TwCMK expression in suspension cell and signally impacted at 24 h. The research provides a basis for further study on the regulation of terpenoid secondary metabolism and biological synthesis.
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