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白桦BplMYB46转录因子的原核表达和重组蛋白的纯化
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作者 国会艳 刘志华 +4 位作者 胡萍 贾园园 张春蕊 杨传平 王玉成 《东北林业大学学报》 CAS CSCD 北大核心 2015年第10期1-5,共5页
从白桦的转录组中发现一个MYB类转录因子Bpl MYB46,将其与拟南芥的MYB类转录因子进行氨基酸序列比对分析,并将其构建到原核表达载体p GEX-4T-2上,获得重组表达载体p GEX-4T-2-Bpl MYB46,然后导入原核表达菌株BL21中,利用异丙基-β-D-硫... 从白桦的转录组中发现一个MYB类转录因子Bpl MYB46,将其与拟南芥的MYB类转录因子进行氨基酸序列比对分析,并将其构建到原核表达载体p GEX-4T-2上,获得重组表达载体p GEX-4T-2-Bpl MYB46,然后导入原核表达菌株BL21中,利用异丙基-β-D-硫代半乳糖(IPTG)诱导重组蛋白的表达。结果显示:Bpl MYB46属于R2R3-MYB类转录因子;在IPTG终浓度为0.4 mmol/L,30℃条件下诱导4 h后p GEX-4T-2-Bpl MYB46原核蛋白表达量最高,主要为包涵体形式;利用尿素梯度透析法和电洗脱法纯化蛋白,发现电洗脱法成功获得浓度和纯度较高的重组蛋白。 展开更多
关键词 白桦 bplmyb46转录因子 原核蛋白 蛋白纯化
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白桦E-box元件的载体构建及与BplMYB46转录因子的互作分析 被引量:2
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作者 王博 王莲萍 +2 位作者 杨春雨 国会艳 魏继承 《生物技术通报》 CAS CSCD 北大核心 2018年第12期110-115,共6页
MYB转录因子是植物中最大的转录因子家族成员之一,主要参与植物次生代谢调控、激素和环境因子的应答,在植物的生长发育中起着至关重要的作用。已有研究发现,E-box的核心序列为CANNTG(N:A/G/C/T),是一类与光响应和苯丙氨酸生物合成途径... MYB转录因子是植物中最大的转录因子家族成员之一,主要参与植物次生代谢调控、激素和环境因子的应答,在植物的生长发育中起着至关重要的作用。已有研究发现,E-box的核心序列为CANNTG(N:A/G/C/T),是一类与光响应和苯丙氨酸生物合成途径相关的元件。在前期研究中,先构建顺式作用元件库,然后以转录因子为中心的酵母单杂交技术筛选发现,白桦的BplMYB46转录因子能够与核心序列为CAAATG的E-box顺式作用元件结合。但是,是否能与E-box的其他核心序列结合还不清楚。本研究将每一种E-box顺式作用元件的核心序列分别进行双链DNA的复性并连接到pHIS2载体上,然后通过酵母单杂交技术筛选与BplMYB46转录因子能够特异结合的E-box顺式作用元件。结果显示,当E-box的核心序列第3个碱基为A/T/C、第四个碱基为A/G/C时,酵母单菌落能在三缺培养基TDO/3AT上生长,表明与BplMYB46转录因子结合的E-box顺式作用元件的特异性序列为CA(A/T/C)(A/G/C)TG。为后续通过BplMYB46转录因子与E-box顺式作用元件的结合来分析BplMYB46转录因子对下游基因的调控,以及为筛选优良下游基因改良白桦的遗传性状奠定数据基础。 展开更多
关键词 bplmyb46转录因子 E-box顺式作用元件 酵母单杂交技术
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Expression profiles of genes regulated by BplMYB46 in Betula platyphylla
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作者 Huiyan Guo Chunrui Zhang +4 位作者 Yanmin Wang Yiming Zhang Yu Zhang Yucheng Wang Chao Wang 《Journal of Forestry Research》 SCIE CAS CSCD 2019年第6期2267-2276,共10页
The transcription factor BplMYB46 has been identified as a regulator of abiotic stress responses and promoter of secondary wall deposition in Betula platyphylla.To investigate the downstream targets of BplMYB46,the ex... The transcription factor BplMYB46 has been identified as a regulator of abiotic stress responses and promoter of secondary wall deposition in Betula platyphylla.To investigate the downstream targets of BplMYB46,the expression profiles of genes in stems from BplMYB46-overexpressing(OE)and BplMYB46-silencing(SE)plants were studied.In OE stems,952 genes were upregulated,and 1469 were downregulated in comparison to SE stems.In a KEGG pathway enrichment analysis of differentially expressed genes(DEGs),1387 differentially expressed genes were annotated for 117 metabolic pathways.DEGs were abundant for metabolic pathway,secondary metabolite biosynthesis,plant hormone signal transduction,phenylpropanoid and flavonoid biosynthesis.DEGs were implicated lignin or cellulose biosynthesis,cell wall modification,xylem development,disease resistance,stress responses,and anthocyanin biosynthesis.These results suggested that BplMYB46 regulates cell wall development and stress resistance by affecting the expression of these genes.Our study further elucidates the mechanism by which BplMYB46 mediates abiotic stress responses and secondary cell wall biosynthesis in birch. 展开更多
关键词 BETULA platyphylla bplmyb46 The GENE EXPRESSION profiles DIFFERENTIALLY EXPRESSED GENES
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Identification of novel cis-elements bound by BplMYB46 involved in abiotic stress responses and secondary wall deposition 被引量:7
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作者 Huiyan Guo Liuqiang Wang +3 位作者 Chuanping Yang Yiming Zhang Chunrui Zhang Chao Wang 《Journal of Integrative Plant Biology》 SCIE CAS CSCD 2018年第10期1000-1014,共15页
Transcription factors (TFs) play vital roles in various biological processes by binding to cis-acting elements to control expressions of their target genes. The MYB TF BplMYB46, from Betula platyphylla, is involved ... Transcription factors (TFs) play vital roles in various biological processes by binding to cis-acting elements to control expressions of their target genes. The MYB TF BplMYB46, from Betula platyphylla, is involved in abiotic stress responses and secondary wall deposition. In the present study, we used a TF-centered yeast onehybrid technology (TF-centered YIH) to identify the cis- acting elements bound by BplMYB46. We screened a shortinsert random library and identified three cis-elements bound by BplMYB46: an E-box (CA(A/T/C)(A/G/C)TG) and two novel motifs, a TO-box (T(GIA)TCG(C/G)) and a GT-box (A(G/T)T(AIC)GT(T/G)C). Chromatin immunoprecipitation (CHIP) and effector-reporter coexpression assays inNicotiana tabacum confirmed binding of BplMYB46 to the TC-box, GT-box, and E-box motifs in the promoters of the phenylalanine ammonia lyase (PAL), peroxidase (POD), and superoxide dismutase (SOD) genes, which function in abiotic stress tolerance and secondary wall biosynthesis. This finding improves our understanding of potential regulatory mechanisms in the response to abiotic stress and secondary wall deposition of BplMYB46 in B. platyphylla. 展开更多
关键词 POD SOD Identification of novel cis-elements bound by bplmyb46 involved in abiotic stress responses and secondary wall deposition cis TCG GUS Figure
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