细辛甲基丁香酚合成相关酶基因的克隆与序列分析  

Cloning and sequence analysis of enzyme genes relevant to biosynthesis of methyl-eugenol in Asarum heterotropoides

作　　者：王鹰 杨悦[1] 李赫 李强[1] 王丽娟[1] 吴秀菊[1] WANG Ying;YANG Yue;LI He;LI Qiang;WANG Li-juan;WU Xiu-ju(College of Life Science, Northeast Agricultural University, Harbin 150030, China)

机构地区：[1]东北农业大学生命科学学院

出　　处：《中草药》2019年第14期3420-3425,共6页Chinese Traditional and Herbal Drugs

基　　金：国家基础科学人才培养基金能力培养与科研训练项目(J1210069);黑龙江省教育厅科学技术研究项目计划(12531030);哈尔滨市科技局科技创新人才研究专项(2012RFLXN003)

摘　　要：目的基于细辛转录组测序结果,克隆甲基丁香酚合成相关酶基因,了解相应的序列信息。方法以细辛叶片为材料,通过RT-PCR扩增得到苯丙氨酸裂解酶(PAL)、肉桂酸-4-羟化酶(C4H)、4-羟基肉桂酰辅酶A连接酶(4CL)、肉桂醇脱氢酶(CAD)等基因的c DNA全长,并进行生物信息学分析。结果 AhPAL、AhC4H、Ah4CL和AhCAD基因的ORF长度分别为2 157、1 278、1 623、1 071 bp,编码718、425、540、356个氨基酸。4种蛋白均具有各自的保守结构域。除Ah C4H外,AhPAL、Ah4CL和AhCAD与已报道的其它物种相应酶的氨基酸序列相似性都较高。结论对细辛甲基丁香酚合成相关酶基因进行克隆和生物信息学分析,为后续基因功能分析及甲基丁香酚生物合成调控机制的解析奠定基础。Objective To clone the enzyme genes related with methyl eugenol synthesis and characterize the corresponding sequence information based on the transcriptome sequencing of Asarum heterotropoides. Methods RT-PCR was performed to obtain the full length cDNA of the phenylalanine lyase (PAL) gene, cinnamic acid-4-hydroxylase (C4H) gene, 4-hydroxycinnamoyl-CoA ligase (4CL), and cinnamyl alcohol dehydrogenase (CAD) genes by using young leaves as materials. The acquired enzyme genes were analyzed by bioinformatics. Results The ORF lengths of AhPAL, AhC4H, Ah4CL, and AhCAD were 2 157, 1 278, 1 623 and 1 071 bp, which respectively encoded 718, 425, 540, and 356 amino acids. Four proteins had respective conserved domains. The amino acid sequences of AhPAL, Ah4CL, and AhCAD were similar to those of other reported species except for AhC4H. Conclusion Four enzyme genes related with methyl-eugenol synthesis in A. heterotropoides were separated and analyzed using bioinformatics method. These results would lay the important foundation for functional analysis of corresponding genes and for elucidating the regulation mechanism of methyl-eugenol biosynthesis.

关 键 词：细辛 甲基丁香酚 基因克隆 序列分析 系统发生分析 

分 类 号：R282.12[医药卫生—中药学]