摘要
目的克隆何首乌Polygonummultiflorum糖基转移酶基因PmUGTs,并对其进行生物信息学以及差异表达分析。方法基于何首乌转录组数据,利用PCR扩增Pm UGTs全长cDNA并进行生物信息学分析;使用autodock4软件进行分子模拟对接;运用q RT-PCR检测基因在不同器官中的表达差异。结果克隆得到4条Pm UGTs,分别命名为Pm UGT1、Pm UGT2、PmUGT3、PmUGT4基因,开放阅读框(ORF)长度分别为1509、1506、1464和1476 bp;编码503、502、488与492个氨基酸;蛋白相对分子质量分别为56 940、54 780、54 350和54 620。Pm UGTs氨基酸序列中含有UGT高度保守的PSPG盒结构域,其二级结构主要由无规则卷曲与α-螺旋组成。进化树分析表明,Pm UGT1、PmUGT2、PmUGT3与拟南芥的UGT蛋白距离较近,Pm UGT4则与蒺藜苜蓿UGT亲缘关系最近。根据autodock4结果可知Pm UGT1、Pm UGT3、Pm UGT2与Pm UGT4蛋白上的残基分别可以与鹰嘴豆芽素A、罗汉果醇、白藜芦醇通过氢键连接。q RT-PCR结果表明,Pm UGTs表达量均在叶中最高。结论为进一步研究何首乌糖基转移酶基因的功能及何首乌中糖苷类化合物的生物合成途径奠定基础。
Objective To clone the Polygonum multiflorum Thunb. glycosyltransferase genes(Pm UGTs) and analyze their bioinformatics and relative expression levels. Methods Based on the transcriptomic data of Polygonum multiflorum Thunb., the Pm UGTs full length c DNA were amplified by PCR and their bioinformatics were analysis. Autodock4 software was used for molecular docking, and qRT-PCR was used to detect Pm UGTs organ-specific expression levels. Results The length of Pm UGT1, PmUGT2, PmUGT3 and PmUGT4 ORF were from 1509, 1506, 1464 and 1476 bp, encoding 503, 502, 488 and 492 amino acids with the molecular mass of 56 940, 54 780, 54 350 and 54 620, respectively. PmUGTs amino acid sequences contain a highly conserved PSPG box domain, and their secondary structures are mainly composed of Random Coil and α-Helix. Phylogenetic analysis showed that PmUGT1, PmUGT2 and PmUGT3 were closed to the Arabidopsis thaliana UGTs, and PmUGT4 was closed to which from Medicago truncatula. According to the autodock4 results, PmUGT1, Pm UGT3, PmUGT2 and PmUGT4 were abled to be hydrogen bonded with biochanin A, mogrol and resveratrol, respectively. Measurement of organ-specific expressions showed that the levels of Pm UGTs leaves expression were the highest. Conclusion This research will laid a foundation for further study on the function of Pm UGTs genes and biosynthesis pathway of P. multiflorum glycosides.
作者
蔡启忠
周良云
刘露
刘长征
王浩
钟磊
杨全
CAI Qi-zhong;ZHOU Liang-yun;LIU Lu;LIU Chang-zheng;WANG Hao;ZHONG Lei;YANG Quan(School of Traditional Chinese Medicine(TCM),Key Laboratory for Production&Development of Cantonese Medicinal Materials Under State Administration of TCM,Guangzhou Comprehensive Experimental StationChinese Materia Medica Industry Technology System,Guangdong Provincial Research Center on Good Agricultural Practice&Comprehensive Agricultural Development Engineering Technology of Cantonese Medicinal Materials,Guangdong Pharmaceutical University,Guangdong Guangzhou 510006,China)
出处
《中草药》
CAS
CSCD
北大核心
2021年第19期6013-6022,共10页
Chinese Traditional and Herbal Drugs
基金
国家重点研发计划项目(2017YFC1700704)
2017年广东省岭南中药材保护资金专项(粤财社[2017]60号)
广东省普通高校青年创新人才类项目(2018KQNCX133)。
关键词
何首乌
糖基转移酶
基因克隆
原核表达
QRT-PCR
Polygonum multiflorum Thunb.
glycosyltransferase
gene cloning
prokaryotic expression
qRT-PCR
