丹参SmbHLH37基因的克隆、亚细胞定位和表达分析  

Cloning,Subcellular Localization and Expression Analysis of SmbHLH37 Gene in Salvia miltiorrhiza

作　　者：苏娇[1] 杨娜[1] 王晓帆[1] 杜堂志 李莎莎[1] 鲁张田子 曹晓燕[1] 

机构地区：[1]陕西师范大学西北濒危药材资源开发国家工程实验室/药用植物资源与天然药物化学教育部重点实验室,西安710062

出　　处：《农业生物技术学报》2017年第6期884-892,共9页Journal of Agricultural Biotechnology

基　　金：国家自然科学基金项目(No.31670299)

摘　　要：碱性螺旋-环-螺旋(basic helix-loop-helix,bHLH)转录因子家族在植物的生长发育、响应非生物胁迫等方面发挥着重要的调控作用。为了进一步丰富丹参(Salvia miltiorrhiza)中该家族成员的研究,寻找新的参与调控丹参次生代谢产物合成相关的bHLH成员,本研究基于丹参转录组和基因组数据库,分别从cDNA和gDNA水平克隆获得SmbHLH37基因序列。该基因序列长1 506 bp,包含1个27 bp的内含子序列,1 479 bp完整的CDS(GenBank登录号:KP257470.1),编码492个氨基酸。Blast P分析结果显示,SmbHLH37与芝麻(Sesamum indicum)等植物的bHLH3同源性较高;SmbHLH37蛋白与拟南芥(Arabidopsis thaliana)bHLH转录因子家族的分子进化树结果显示,SmbHLH37与拟南芥bHLH转录因子At JAM3(A.thaliana jasmonate-associated MYC2-like3)亲缘关系最近。亚细胞定位结果显示,SmbHLH37蛋白主要定位在细胞核中。qRT-PCR分析结果显示,SmbHLH37在叶中的表达量最高,花中最低;用脱落酸(abscisic acid,ABA)和机械诱导处理后,该基因的表达短时间呈上调趋势;同时,SmbHLH37在Sm MYC2过表达株系OE-8和OE-12中的表达量显著上调(P<0.05)。研究结果为进一步开展SmbHLH37功能研究提供了基础资料。Basic helix-loop-helix （bHLH） transcription factors play important roles in regulating plant growth-development and abiotic stress response. In order to enrich the research content of bHLH transcription factors and find new bHLH transcription factor in regulating secondary metabolite in Salvia miltiorrhiza, SmbHLH37 was cloned based on gDNA and cDNA. The gene sequence of SmbHLH37 was 1 506 bp with one intron of 27 bp, and 1 479 bp complete CDS （GenBank accession No. KP257470.1） encoding 492 amino acids. BlastP result showed that SmbHLH37 protein had high homology with bHLH3 in Sesamum indicum etc. Amino acid sequence analysis showed that SmbHLH37 contained a HLH domain and motif 7. Molecular evolutionary tree of SmbHLH37 protein and all the bHLH transcription factors in Arabidopsis thaliana indicated that SmbHLH37 protein was the closest relative to AtJAM3 （A. thaliana jasmonate-associated MYC2-1ike 3）. Subcellular localization showed that SmbHLH37 protein was mainly localized in the nucleus. The expression pattern disclosed that the expression level of SmbHLH37 was the highest in leaves and the lowest in flowers, qRT- PCR analysis indicated that expression of SmbHLH37 could increase in a short time with 0.1 mmol/L abscisic acid （ABA） and wounding treatment. SmMYC2 and SmbHLH37 expression levels in SmMYC2 overexpressing transgenic S. miltiorrhiza lines OE-8 and OE-12 were detected, and the results showed SmMYC2 expression levels in OE-8 and OE-12 significantly or extremely significantly increased （P〈 0.05 or P〈0.001）. Meanwhile, the expression levels of SmbHLH37 in OE-8 and OE-12 were similar with SmMYC2 expression. The present data has laid a foundation for studying the function of SmbHLH37 in the future.

关 键 词：丹参 碱性螺旋-环-螺旋(bHLH) 生物信息学分析 表达分析 亚细胞定位 

分 类 号：S184[农业科学—农业基础科学]