Prokaryotic Expression and Purification of <i>JsMYB</i>305 Transcription Factor in <i>Jasminum sambac</i>

WAN Chao; ZHANG Yue; HU Li; WU Binghua; YUAN Yuan

doi:10.19303/j.issn.1008-0384.2022.002.005

Volume 37 Issue 2

Feb. 2022

Turn off MathJax

Article Contents

Article Navigation > Fujian Journal of Agricultural Sciences > 2022 > 37(2): 164-169

WAN C, ZHANG Y, HU L, et al. Prokaryotic Expression and Purification of JsMYB305 Transcription Factor in Jasminum sambac [J]. Fujian Journal of Agricultural Sciences，2022，37（2）：164−169 doi: 10.19303/j.issn.1008-0384.2022.002.005

Citation:

WAN C, ZHANG Y, HU L, et al. Prokaryotic Expression and Purification of JsMYB305 Transcription Factor in Jasminum sambac [J]. Fujian Journal of Agricultural Sciences，2022，37（2）：164−169 doi: 10.19303/j.issn.1008-0384.2022.002.005

Citation:

PDF( 808 KB)

Prokaryotic Expression and Purification of JsMYB305 Transcription Factor in Jasminum sambac

doi: 10.19303/j.issn.1008-0384.2022.002.005

College of Horticulture, Fujian Agriculture and Forestry University, Fuzhou, Fujian　350002, China

Received Date: 2021-10-21
Rev Recd Date: 2022-01-11

Available Online: 2022-02-07

Publish Date: 2022-02-25

Abstract

Abstract

Objective The recombinant protein of fragrance-regulating transcription factor JsMYB305 was obtained in preparation for studies on the molecular mechanism of terpenoid-regulation and interacting proteins of the gene in Jasminum sambac. Methods Coding sequence of JsMYB305 was constructed into prokaryotic expression vector pGEX-4T-1 by enzyme digestion and ligation, then, transformed into the BL21 (DE3) expressing strain. The protein expression was induced by IPTG, and the recombinant protein separated and purified using GST affinity resin followed by western blot for positive identification. Result After induction by 0.2 mmol·L⁻¹ IPTG, the recombinant protein was purified at the optimized temperature of 28 ℃ for 4 h and eluted with 20 mmol·L⁻¹ GSH for purification. The western blot confirmed the protein weight to be as expected. Conclusion The recombinant protein of JsMYB305 was successfully obtained to pave the way for further studies on the search for the interacting proteins by GST-pull down technique and the binding of the gene to specific promoter sites by EMSA.
- Jasminum sambac,
- MYB transcription factor,
- prokaryotic expression,
- protein purification

FullText(HTML)

References(28)

References

[1]	朱建新, 程福建, 杨江帆. 茉莉花茶香气影响因素研究进展 [J]. 中国茶叶, 2021, 43(1):40−43. doi: 10.3969/j.issn.1000-3150.2021.01.007 ZHU J X, CHENG F J, YANG J F. Research progress on factors influencing aroma of jasmine tea [J]. China Tea, 2021, 43(1): 40−43.(in Chinese) doi: 10.3969/j.issn.1000-3150.2021.01.007
[2]	BERA P, KOTAMREDDY J N R, SAMANTA T, et al. Inter-specific variation in headspace scent volatiles composition of four commercially cultivated jasmine flowers [J]. Natural Product Research, 2015, 29(14): 1328−1335. doi: 10.1080/14786419.2014.1000319
[3]	BERA P, MUKHERJEE C, MITRA A. Enzymatic production and emission of floral scent volatiles in Jasminum sambac [J]. Plant Science, 2017, 256: 25−38. doi: 10.1016/j.plantsci.2016.11.013
[4]	SPITZER-RIMON B, MARHEVKA E, BARKAI O, et al. EOBII, a gene encoding a flower-specific regulator of phenylpropanoid volatiles' biosynthesis in Petunia [J]. The Plant Cell, 2010, 22(6): 1961−1976. doi: 10.1105/tpc.109.067280
[5]	SPITZER-RIMON B, FARHI M, ALBO B, et al. The R2R3-MYB-like regulatory factor EOBI, acting downstream of EOBII, regulates scent production by activating ODO1 and structural scent-related genes in Petunia [J]. The Plant Cell, 2012, 24(12): 5089−5105.
[6]	VAN MOERKERCKE A, HARING M A, SCHUURINK R C. The transcription factor emission of benzenoids ii activates the MYB odorant1 promoter at a MYB binding site specific for fragrant petunias [J]. The Plant Journal:for Cell and Molecular Biology, 2011, 67(5): 917−928. doi: 10.1111/j.1365-313X.2011.04644.x
[7]	MEDINA-PUCHE L, MOLINA-HIDALGO F J, BOERSMA M, et al. An R2R3-MYB transcription factor regulates eugenol production in ripe strawberry fruit receptacles [J]. Plant Physiology, 2015, 168(2): 598−614. doi: 10.1104/pp.114.252908
[8]	JIAN W, CAO H H, YUAN S, et al. SlMYB75, an MYB-type transcription factor, promotes anthocyanin accumulation and enhances volatile aroma production in tomato fruits [J]. Horticulture Research, 2019, 6: 22. doi: 10.1038/s41438-018-0098-y
[9]	孙君, 陈雪津, 陈笛, 等. 茉莉花JsPAL基因及其启动子克隆与表达分析 [J]. 西北植物学报, 2020, 40(6):949−956. SUN J, CHEN X J, CHEN D, et al. Cloning and expression analysis of phenylalanine ammonia-lyase gene from Jasminum sambac and isolation of its promoter [J]. Acta Botanica Boreali-Occidentalia Sinica, 2020, 40(6): 949−956.(in Chinese)
[10]	陈笛, 王鹏杰, 郑玉成, 等. 茉莉花MVD基因及其启动子的克隆与表达 [J]. 福建农林大学学报(自然科学版), 2019, 48(3):309−315. CHEN D, WANG P J, ZHENG Y C, et al. Cloning and expression of MVD gene and its promoter in Jasminum sambac [J]. Journal of Fujian Agriculture and Forestry University (Natural Science Edition), 2019, 48(3): 309−315.(in Chinese)
[11]	陈笛, 陈雪津, 郭永春, 等. 茉莉花芳樟醇生物合成关键基因的克隆与表达分析[J]. 西北植物学报, 2019, 39（8）: 1344−1352. CHEN D, CHEN X J, GUO Y C, et al. Cloning and expression analysis of JsNEL/LINS from Jasminum sambac[J]. Acta Botanica Boreali-Occidentalia Sinica, 2019, 39(8): 1344−1352.（in Chinese）
[12]	崔萌, 刘志钦, 叶乃兴, 等. 茉莉花香气相关基因JsGDS启动子的克隆及功能分析 [J]. 分子植物育种, 2021, 19(2):441−447. CUI M, LIU Z Q, YE N X, et al. Cloning and functional analysis of the aroma-related gene JsGDS promoter from Jasminum sambac [J]. Molecular Plant Breeding, 2021, 19(2): 441−447.(in Chinese)
[13]	陈桂信, 俞滢, 刘雨轩, 等. 茉莉香气释放相关基因分离及JsBEBT的克隆与表达分析 [J]. 分子植物育种, 2021, 19(20):6662−6670. CHEN G X, YU Y, LIU Y X, et al. Isolation of related genes cloning and expression analysis of JsBEBT in jasmine aroma [J]. Molecular Plant Breeding, 2021, 19(20): 6662−6670.(in Chinese)
[14]	陈笛, 郑玉成, 林浥, 等. 茉莉花MK基因及启动子克隆与表达分析 [J]. 分子植物育种, 2020, 18(3):772−779. CHEN D, ZHENG Y C, LIN Y, et al. Cloning and expression analysis of mevalonate kinase gene and its promoter from Jasminum sambac [J]. Molecular Plant Breeding, 2020, 18(3): 772−779.(in Chinese)
[15]	崔萌. 茉莉花香气合成相关基因JsGDS启动子的分离与活性分析[D]. 福州: 福建农林大学, 2020. CUI M. Isolation and activity analysis of promoter of JsGDS gene associated with aroma synthesis from Jasminum sambac[D]. Fuzhou: Fujian Agriculture and Forestry University, 2020. (in Chinese)
[16]	孙君, 陈桂信, 叶乃兴, 等. 茉莉花香气相关基因JsDXS及其启动子的克隆与表达分析 [J]. 园艺学报, 2014, 41(6):1236−1244. SUN J, CHEN G X, YE N X, et al. Cloning and expression analysis of deoxyoxylulose-5-phosphate synthase gene related to aroma from Jasminum sambac and isolation of its promoter [J]. Acta Horticulturae Sinica, 2014, 41(6): 1236−1244.(in Chinese)
[17]	张月, 袁媛, 何弦, 等. 茉莉花JsMYB108和JsMYB305基因的克隆及其对TPS基因的激活作用 [J]. 热带作物学报, 2021, 42(6):1539−1548. doi: 10.3969/j.issn.1000-2561.2021.06.005 ZHANG Y, YUAN Y, HE X, et al. Cloning of JsMYB108 and JsMYB305 and analysis of their activation on TPS gene in Jasminum sambac [J]. Chinese Journal of Tropical Crops, 2021, 42(6): 1539−1548.(in Chinese) doi: 10.3969/j.issn.1000-2561.2021.06.005
[18]	张磊, 唐永凯, 李红霞, 等. 促进原核表达蛋白可溶性的研究进展 [J]. 中国生物工程杂志, 2021, 41(S1):138−149. ZHANG L, TANG Y K, LI H X, et al. Advances in promoting solubility of prokaryotic expressed proteins [J]. China Biotechnology, 2021, 41(S1): 138−149.(in Chinese)
[19]	邱淑彬, 荆韧威, 郭正隆, 等. 两种不同的原核表达载体蛋白表达及纯化的比较 [J]. 基因组学与应用生物学, 2020, 39(9):4136−4144. QIU S B, JING R W, GUO Z L, et al. Comparison of two different prokaryotic expression vectors in protein expression and purification [J]. Genomics and Applied Biology, 2020, 39(9): 4136−4144.(in Chinese)
[20]	黄传臻, 刘香利, 曹汝菲, 等. 小麦CWI-B1的原核表达、纯化与多克隆抗体制备 [J]. 农业生物技术学报, 2017, 25(7):1102−1110. HUANG C Z, LIU X L, CAO R F, et al. Prokaryotic expression, purification and preparation of polyclonal antibody for wheat (Triticum aestivum) CWI-B1 [J]. Journal of Agricultural Biotechnology, 2017, 25(7): 1102−1110.(in Chinese)
[21]	陈明, 陈文静. 几种微生物表达系统的比较 [J]. 安徽农学通报(上半月刊), 2011, 17(3):68−71. CHEN M, CHEN W J. Comparisons of some microbial expression systems [J]. Anhui Agricultural Science Bulletin, 2011, 17(3): 68−71.(in Chinese)
[22]	胡京蕊, 沈金宝, 李晶岚, 等. 拟南芥耐盐相关基因AtSTK原核表达载体的构建及表达 [J]. 华北农学报, 2013, 28(2):38−41. doi: 10.3969/j.issn.1000-7091.2013.02.007 HU J R, SHEN J B, LI J L, et al. Prokaryotic expression vector construction and expression of tolerance related gene AtSTK in Arabidopsis [J]. Acta Agriculturae Boreali-Sinica, 2013, 28(2): 38−41.(in Chinese) doi: 10.3969/j.issn.1000-7091.2013.02.007
[23]	孙伟, 任家鑫, 腾峰, 等. ZmAGO18b的原核表达载体的构建与表达 [J]. 基因组学与应用生物学, 2020, 39(12):5647−5651. SUN W, REN J X, TENG F, et al. Construction and expression of a prokaryotic expression vector of ZmAGO18b [J]. Genomics and Applied Biology, 2020, 39(12): 5647−5651.(in Chinese)
[24]	HARPER S, SPEICHER D W. Purification of proteins fused to glutathione S-transferase [J]. Methods in Molecular Biology (Clifton, N J ), 2011, 681: 259−280.
[25]	朱炳森. 玉米转录因子ZmYY1、ZmYY2与RNA结合蛋白ZmRBM25互作调控ZmABR1表达的研究[D]. 泰安: 山东农业大学, 2020. ZHU B S. Maize transcription factors ZmYY1, ZmYY2 interact with RNA-binding protein ZmRBM25 to regulate ZmABR1 expression[D]. Taian: Shandong Agricultural University, 2020. (in Chinese)
[26]	王意程. 油菜素内酯调控红肉苹果类黄酮合成的机理研究[D]. 泰安: 山东农业大学, 2021. WANG Y C. Mechanism of brassinosteroid on regulation flavonoid biosynthesis in red-fleshed apple[D]. Taian: Shandong Agricultural University, 2021. (in Chinese)
[27]	赵杰, 王兵, 骆梅, 等. GST-pull down技术筛选毛白杨天冬氨酸蛋白酶PtoAED3互作蛋白 [J]. 北京林业大学学报, 2021, 43(5):64−74. doi: 10.12171/j.1000-1522.20200365 ZHAO J, WANG B, LUO M, et al. Identification of aspartic acid protease PtoAED3-interacting proteins through GST pull-down assays in Populus tomentosa [J]. Journal of Beijing Forestry University, 2021, 43(5): 64−74.(in Chinese) doi: 10.12171/j.1000-1522.20200365
[28]	窦万福, 祁静静, 胡安华, 等. GST pull-down联合LC-MS/MS筛选柑橘抗溃疡病转录因子CsBZIP40的互作蛋白 [J]. 中国农业科学, 2019, 52(13):2243−2255. doi: 10.3864/j.issn.0578-1752.2019.13.005 DOU W F, QI J J, HU A H, et al. Screening of interacting proteins of anti-canker transcription factor CsBZIP40 in Citrus by GST pull-down combined with LC-MS/MS [J]. Scientia Agricultura Sinica, 2019, 52(13): 2243−2255.(in Chinese) doi: 10.3864/j.issn.0578-1752.2019.13.005