| 摘要: |
| 该研究在夏枯草转录组测序的基础上设计特异引物,采用逆转录PCR技术获得该基因的全长核苷酸序列,并进行生物信息学分析,采用qRT-PCR法分析PvDXS在夏枯草不同组织及不同外源性物质诱导下的表达量。结果表明:克隆得到的PvDXS基因开放阅读框2 181 bp,编码726个氨基酸,理论分子量为78 040.47 D,等电点为6.75,PvDXS蛋白具有Transketolase_C结构域和Transket_pyr结构域,系统进化树结果表明,PvDXS蛋白与丹参、长春花的DXS(SmDXS2、CrDXS2)亲缘关系较近,推测PvDXS属于第Ⅱ类DXS蛋白。qRT-PCR分析表明,PvDXS基因在叶中表达量高于果穗及茎。对果穗施加7种外源性物质处理24 h后,GA3处理组该基因表达量升高,其他6种外源性物质处理后表达量均降低,其中CaCl2、SNP、SA处理后该基因的表达量显著降低。PvDXS基因在不同组织中表达量差异较大,且受外源物质诱导表达。这为进一步研究PvDXS基因对夏枯草萜类成分合成途径中的功能及表达调控奠定基础。 |
| 关键词: 夏枯草, PvDXS基因, 基因克隆, 表达分析 |
| DOI：10.11931/guihaia.gxzw201811025 |
| 分类号:Q943.2 |
| 文章编号:1000-3142(2019)12-1619-09 |
| Fund project:国家自然科学基金(81603232); 国家重点研发计划项目(2017YFC1702800); 河南中医学院博士科研基金(BSJJ2015-13)[Supported by the National Natural Science Foundation of China(81603232); the National Key Research and Development Program of China(2017YFC1702800); Doctoral Research Fund of Henan University of Traditional Chinese Medicine(BSJJ2015-13)]。 |
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| Cloning and expression analysis of PvDXS gene from Prunella vulgaris |
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LI Lu1, DONG Chengming1,2, ZHANG Mengjia1, ZHU Yunhao1,2*
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1. School of Pharmacy, Henan University of Traditional Chinese Medicine, Zhengzhou 450046, China;2. Collaborative Innovation Center for
Respiratory Disease Diagnosis and Treatment &3.Chinese Medicine Development of Henan Province, Co-construction Collaborative Innovation
Center for Chinese Medicine and Respiratory Diseases by Henan &4.Education Ministry of PR. China, Zhengzhou 450046, China
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| Abstract: |
| Specific primers were designed on the basis of transcriptome sequencing of Prunella vulgaris. The full-length nucleotide sequence of PvDXS was obtained by reverse transcription PCR and the bioinformatics analysis of the gene was conducted. The expression levels of PvDXS in different tissues and exogenous substances were detected by real-time quantitative PCR. The results showed the cDNA sequence of PvDXS contained the open reading frame which had 2 181 bp and encoded a predicted protein of 726 amino acids with a theoretical molecular weight of 78 040.47 D and a isoelectric point of 6.75. The protein had Transketolase_C domain and Transket_pyr domain. Phylogenetic tree results showed that PvDXS protein was closely related to DXS(SmDXS2, CrDXS2)from Salvia miltiorrhiza and Catharanthus roseus, and it was inferred that PvDXS belonged to the Class Ⅱ DXS protein type. Tissue expression pattern analysis revealed that PvDXS gene in leaves was higher than that in ears and stems. After treated with seven exogenous substances for 24 h, the expression of the gene increased in GA3 treatment group and decreased after treatment with the others. The expression level of the gene decreased significantly after CaCl2, SNP and SA treatments. The expressions of PvDXS were variant in different tissues and varied greatly after treatment of exogenous substances, which laid a foundation for further study on the function and expression regulation of PvDXS in the synthesis pathway of terpenoid components of Prunella vulgaris. |
| Key words: Prunella vulgaris, 1-deoxy-D-xylulose 5-phosphate synthase gene, gene clone, expression analysis |
