摘要
WRKY转录因子是植物特有的一类转录因子,在植物多种生理过程中发挥重要的作用。本研究以茅苍术转录组数据为依据,对茅苍术AlWRKY基因与AlTPS基因的FPKM值进行相关性分析,结合分析结果筛选出与AlTPS1、AlTPS6基因表达量具有显著正相关性且FPKM值相对较高的候选基因AlWRKY65,对该候选基因进行克隆,获得AlWRKY65基因的开放阅读框(ORF),分析其编码蛋白的相关信息并进行基因表达研究。研究结果表明,AlWRKY65包含一个681bp的开放阅读框,编码226个氨基酸。AlWRKY65基因编码的氨基酸序列与向日葵HaWRKY65、莴苣LsWRKY65等植物基因编码的氨基酸序列具有较高同源性;该蛋白具有1个WRKYGQK保守结构域,属于WRKY基因家族的第IIe组;系统进化树分析显示,茅苍术AlWRKY65与洋蓟CcWRKY65蛋白亲缘关系较近;运用实时荧光定量PCR检测AlWRKY65基因在2个产地不同组织中的表达水平,结果表明,不同产地的茅苍术AlWRKY65基因具有组织表达差异性,均在叶片中表达量较高;在茉莉酸甲酯(MeJA)诱导处理48 h内,AlWRKY65基因的表达量下调;亚细胞定位和转录自激活活性分析显示,AlWRKY65定位于细胞核,不具有自激活活性。这些结果为进一步研究AlWRKY65在茅苍术中的生物学功能提供参考。
WRKY transcription factor is a type of transcription factor unique to plants and plays an important role in various physiological processes of plants.This study is based on the transcriptome data of Atractylodes lancea,the correlation between the FPKM values of the AlWRKY gene and the AlTPS gene of Atractylodes lancea was analyzed.Combined with the analysis results,the candidate gene AlWRKY65 with a significant positive correlation with the expression levels of AlTPS1 and AlTPS6 genes and a relatively high FPKM value was screened.The candidate gene was cloned to obtain the open reading frame ORF of the AlWRKY65 gene,and the related information of its encoded protein was analyzed and the gene expression was studied.The results showed that AlWRKY65 contains a 681 bp open reading frame and encoding 226 amino acids.Through amino acid sequence homology analysis,it was found that AlWRKY65 amino acid sequence had high homology with several plants such as HaWRKY65 and LsWRKY65;AlWRKY65 protein had a typical WRKYGQK domain,belonging to IIe subgroup of WRKY transcription factor family;phylogenetic analysis indicated that AlWRKY65 protein had the higher homology with CcWRKY65 protein;the expression of AlWRKY65 gene in different tissues of two producing areas of Atractylodes lancea were assayed via real-time fluorescence quantitative PCR,and the results showed that all of them were highly expressed in leaves and also has tissue differences.The expression level of AlWRKY65 gene was down-regulated within 48 h of methyl jasmonate(MeJA)induction;subcellular localization and transcriptional activation assay suggested that AlWRKY65 was located in the nucleus and had no transcriptional activation activity.This study provides a reference for further elucidating the biological function of AlWRKY65 in Atractylodes lancea.
作者
管凤雅
刘巍玮
迟凯文
曾凯玲
谢晋
查良平
GUAN Feng-ya;LIU Wei-wei;CHI Kai-wen;ZENG Kai-ling;XIE Jin;ZHA Liang-ping(School of Pharmacy,Anhui University of Chinese Medicine,Hefei 230012,China;School of Pharmacy,Anhui Medical University,Hefei 230032,China)
出处
《药学学报》
CAS
CSCD
北大核心
2024年第5期1494-1502,共9页
Acta Pharmaceutica Sinica
基金
国家自然科学基金面上项目(82073957)
安徽省自然科学基金优青项目(2208085Y30)
安徽省高校杰出青年科研项目(2023AH020036)
安徽省高校优秀青年人才支持计划重点项目(gxyqZD2022051)
安徽高校自然科学研究重点项目(2023AH050634)
中华中医药学会青年人才托举工程项目(CACM-2023-QNRC2-B23)。
关键词
茅苍术
WRKY转录因子
基因克隆
表达分析
亚细胞定位
Atractylodes lancea
WRKY transcription factor
gene cloning
expression analysis
subcellular localization
