Article(id=1304414879004258769, tenantId=1146029695717560320, journalId=1302319053441957962, issueId=1304414858296971266, articleNumber=null, orderNo=null, doi=10.7501/j.issn.0253-2670.2026.06.021, pmid=null, cstr=null, oa=null, hot=0, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=null, receivedDate=1762099200000, receivedDateStr=2025-11-03, revisedDate=null, revisedDateStr=null, acceptedDate=null, acceptedDateStr=null, onlineDate=1788926342010, onlineDateStr=2026-09-09, pubDate=null, pubDateStr=null, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1788926342010, onlineIssueDateStr=2026-09-09, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1788926342010, creator=13701087609, updateTime=1788926342010, updator=13701087609, issue=Issue{id=1304414858296971266, tenantId=1146029695717560320, journalId=1302319053441957962, year='2026', volume='57', issue='6', pageStart='2009', pageEnd='2444', issueExtLink='null', onlineDate='null', pubDate='1774627200000', pubDateStr='2026-03-28', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=null, createTime=1788926337074, creator='13701087609', updateTime=1788926665348, updator='13701087609', preIssue=null, nextIssue=null, articleTotal=null, ext={EN=IssueExt(id=1304416235240841997, tenantId=1146029695717560320, journalId=1302319053441957962, issueId=1304414858296971266, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1304416235240841998, tenantId=1146029695717560320, journalId=1302319053441957962, issueId=1304414858296971266, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null, downloadFileDto=null}, startPage=2265, endPage=2277, ext={EN=ArticleExt(id=1304414879566295510, articleId=1304414879004258769, tenantId=1146029695717560320, journalId=1302319053441957962, language=EN, title=Cloning, prokaryotic expression, and activity analysis of two MAPK genes in Aquilaria sinensis, columnId=null, journalTitle=Chinese Traditional and Herbal Drugs, columnName=null, runingTitle=null, highlight=null, articleAbstract=ObjectiveAsMAPK1 and AsMAPK2 are two mitogen-activated protein kinase (MAPK) genes with unknown functions in Aquilaria sinensis. To investigate their fundamental characteristics and potential functions in wound-induced agarwood formation, in this study, we performed cloning, expression analysis, and preliminary in vitro functional assays on the two genes. Methods Using cDNA from subcultured A. sinensis callus as the template, the coding sequences of AsMAPK1 and AsMAPK2 were obtained by PCR amplification. Bioinformatics analysis was performed using relevant online tools and softwares. The prokaryotic expression vectors pGEX4T-1-AsMAPK1/2 were constructed, transformed into E. coli BL21 for expression, and induction conditions were optimized. The AsMAPK1-GFP and AsMAPK2-GFP fusion expression vectors were constructed and transformed into Arabidopsis thaliana protoplasts to observe their subcellular localization using confocal microscopy. The kinase activity of the purified proteins was detected using an in vitro phosphorylation assay. Results The AsMAPK1 (1 188 bp) and AsMAPK2 (1 128 bp) genes were successfully cloned. Bioinformatics analysis revealed that both encoded proteins contain typical kinase domains and belong to the MAPK family. Phylogenetic analysis indicated that AsMAPK1 is most closely related to the AtMAPK6 homolog in A. thaliana and belongs to Group A, while AsMAPK2 clusters with Group B AtMAPK13. SDS-PAGE detection showed successful induction of soluble recombinant proteins of approximately 71 000 and 69 000. Subcellular localization results demonstrated that both AsMAPK1-GFP and AsMAPK2-GFP are localized in the cytoplasm and nucleus. Further kinase activity analysis confirmed that both AsMAPK1 and AsMAPK2 undergo autophosphorylation and can phosphorylate the universal substrate MBP. Conclusion This study confirms that AsMAPK1 and AsMAPK2 are active protein kinases localized in both the cytoplasm and nucleus. The cloning, expression analysis, and kinase activity detection of the two AsMAPKs provide a foundation for further elucidating their roles in the signal transduction mechanism underlying wound-induced agarwood formation in A. sinensis., authors=YANG Shuwen, RONG Mei, WANG Siyu, SUN Yuxuan, XU Yanhong, WEI Jianhe, authorsList=YANG Shuwen, RONG Mei, WANG Siyu, SUN Yuxuan, XU Yanhong, WEI Jianhe, authorCompany=null, correspAuthors=null, authorNote=null, correspAuthorsNote=null, copyrightStatement=null, copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=null, magXml=null, pdfUrl=null, pdf=null, pdfFileSize=null, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=null, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=null, mapNumber=null, fund=null), CN=ArticleExt(id=1304414879465632213, articleId=1304414879004258769, tenantId=1146029695717560320, journalId=1302319053441957962, language=CN, title=白木香中2个MAPK基因的克隆、原核表达及活性分析, columnId=1304140203967410414, journalTitle=中草药, columnName=药材与资源, runingTitle=null, highlight=null, articleAbstract=目的AsMAPK1和AsMAPK2是白木香Aquilaria sinensis中2个功能未知的丝裂原活化蛋白激酶基因，为探究其基本特性及其在伤害诱导沉香形成中潜在功能，对其进行克隆、表达分析及初步体外功能检测。方法 以课题组继代培养的白木香愈伤组织cDNA为模板，PCR扩增获得AsMAPK1和AsMAPK2的编码序列；利用相关在线工具及软件进行生物信息学分析；构建pGEX4T-1-AsMAPK1/2原核表达载体，转化大肠杆菌BL21表达并对诱导条件进行优化；构建AsMAPK1-GFP和AsMAPK2-GFP融合表达载体，转化拟南芥原生质体后利用共聚焦显微镜观察其亚细胞定位；采用体外磷酸化方法检测纯化蛋白的激酶活性。结果 成功克隆了AsMAPK1（1 188 bp）和AsMAPK2（1 128 bp）基因。生物信息学分析显示，二者编码的蛋白均含有典型的激酶结构域，属于MAPK家族成员。系统进化分析表明，AsMAPK1与拟南芥中的AtMAPK6同源蛋白亲缘关系最近，属于A组，而AsMAPK2则与B组的AtMAPK13聚为一支。SDS-PAGE检测显示，成功诱导表达了约71 000和69 000的可溶性重组蛋白。亚细胞定位结果表明，AsMAPK1-GFP和AsMAPK2-GFP都同时定位于细胞质和细胞核。进一步的激酶活性分析证实，AsMAPK1和AsMAPK2均能发生自磷酸化，并能磷酸化通用底物MBP。结论 证实AsMAPK1和AsMAPK2是同时定位于细胞质和细胞核、具有活性的蛋白激酶，通过对这2个基因的克隆、表达与分析，为深入解析其参与白木香伤害诱导形成沉香的信号传导机制奠定基础。, authors=杨舒雯1, 戎梅1, 王斯雨1, 孙雨萱1, 徐艳红1, 魏建和1,2, authorsList=杨舒雯, 戎梅, 王斯雨, 孙雨萱, 徐艳红, 魏建和, authorCompany=1 中国医学科学院 北京协和医学院药用植物研究所 中草药物质基础与资源利用教育部重点实验室 濒危药材繁育国家工程实验室, 北京 100193; 2 中国医学科学院 北京协和医学院药用植物研究所海南分所 海南省南药资源保护与开发重点实验室 国家中医药管理局沉香可持续利用重点研究室, 海南 海口 570311, correspAuthors=null, authorNote=杨舒雯: 杨舒雯,硕士研究生,研究方向为药用植物次生代谢调控机制。E-mail:s2023009040@student.pumc.edu.cn, correspAuthorsNote=null, copyrightStatement=null, copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=null, magXml=null, pdfUrl=null, pdf=iofy/LRtbJuNSAyv5DwAhg==, pdfFileSize=1718194, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=null, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=null, mapNumber=null, fund=国家自然科学基金项目 (82173925); 中国医学科学院医学与健康科技创新工程—重大协同创新项目 (2021-I2M-1-032))}, authors=null, keywords=[Keyword(id=1304414879721484759, tenantId=1146029695717560320, journalId=1302319053441957962, articleId=1304414879004258769, language=CN, orderNo=1, keyword=白木香), Keyword(id=1304414879796982234, tenantId=1146029695717560320, journalId=1302319053441957962, articleId=1304414879004258769, language=CN, orderNo=2, keyword=丝裂原活化蛋白激酶（MAPK）), Keyword(id=1304414879885062621, tenantId=1146029695717560320, journalId=1302319053441957962, articleId=1304414879004258769, language=CN, orderNo=3, keyword=克隆), Keyword(id=1304414879968948702, tenantId=1146029695717560320, journalId=1302319053441957962, articleId=1304414879004258769, language=CN, orderNo=4, keyword=生信分析), Keyword(id=1304414880031863263, tenantId=1146029695717560320, journalId=1302319053441957962, articleId=1304414879004258769, language=CN, orderNo=5, keyword=原核表达), Keyword(id=1304414880166080992, tenantId=1146029695717560320, journalId=1302319053441957962, articleId=1304414879004258769, language=EN, orderNo=1, keyword=Aquilaria sinensis (Lour.) 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provinceName=null, collectFlag=false, interPubPlatform=, interPubPlatformUrl=null), detailUrlCn=https://castjournals.cast.org.cn/joweb/zcy/CN/10.7501/j.issn.0253-2670.2026.06.021, detailUrlEn=https://castjournals.cast.org.cn/joweb/zcy/EN/10.7501/j.issn.0253-2670.2026.06.021, pdfUrlCn=https://castjournals.cast.org.cn/joweb/zcy/CN/PDF/10.7501/j.issn.0253-2670.2026.06.021, pdfUrlEn=https://castjournals.cast.org.cn/joweb/zcy/EN/PDF/10.7501/j.issn.0253-2670.2026.06.021, aliStartDate=null, aliEndDate=null, collectionFlag=false, citedCount=null, citedUrl=null, previewStatus=0, delFlag=0, hasFullText=0, orderTime=1788926342010, fullTextJson=null, articleText=null, reference=李林海, 寿海洋, 马清温. 土沉香(瑞香科)的地理分布研究[J]. 安徽农业科学, 2012, 40(17): 9254-9256. 张争, 杨云, 魏建和, 等. 白木香结香机制研究进展及其防御反应诱导结香假说[J]. 中草药, 2010, 41(1): 156-159. Li W, Chen H Q, Wang H, et al. Natural products in agarwood and Aquilaria plants: Chemistry, biological activities and biosynthesis[J]. Nat Prod Rep, 2021, 38(3): 528-565. Wang S, Yu Z X, Wang C H, et al. Chemical constituents and pharmacological activity of agarwood and Aquilaria plants[J]. Molecules, 2018, 23(2): 342. 雷胄熙, 吴泽青, 林励, 等. 生物诱导法所产不同部位沉香挥发油成分分析[J]. 中药新药与临床药理, 2016, 27(1): 106-113. 林峰, 戴好富, 王辉, 等. 两批接菌法所产沉香挥发油化学成分的气相色谱-质谱联用分析[J]. 时珍国医国药, 2010, 21(8): 1901-1902. Liu Y Y, Chen H Q, Yang Y, et al. Whole-tree agarwood-inducing technique: An efficient novel technique for producing high-quality agarwood in cultivated Aquilaria sinensis trees[J]. Molecules, 2013, 18(3): 3086-3106. Jiang M, Zhang Y Z, Li P, et al. Mitogen-activated protein kinase and substrate identification in plant growth and development[J]. Int J Mol Sci, 2022, 23(5): 2744. Xu J, Zhang S Q. Mitogen-activated protein kinase cascades in signaling plant growth and development[J]. Trends Plant Sci, 2015, 20(1): 56-64. Zhang M M, Zhang S Q. Mitogen-activated protein kinase cascades in plant signaling[J]. J Integr Plant Biol, 2022, 64(2): 301-341. Mao G H, Meng X Z, Liu Y D, et al. Phosphorylation of a WRKY transcription factor by two pathogen-responsive MAPKs drives phytoalexin biosynthesis in Arabidopsis[J]. Plant Cell, 2011, 23(4): 1639-1653. Meng X Z, Xu J, He Y X, et al. Phosphorylation of an ERF transcription factor by Arabidopsis MPK3/MPK6 regulates plant defense gene induction and fungal resistance[J]. Plant Cell, 2013, 25(3): 1126-1142. Kishi-Kaboshi M, Seo S, Takahashi A, et al. The MAMP-responsive MYB transcription factors MYB30, MYB55 and MYB110 activate the HCAA synthesis pathway and enhance immunity in rice[J]. Plant Cell Physiol, 2018, 59(5): 903-915. Zhang D, Jiang S, Pan J, et al. The overexpression of a maize mitogen-activated protein kinase gene (ZmMPK5) confers salt stress tolerance and induces defence responses in tobacco[J]. Plant Biol, 2014, 16(3): 558-570. Long L, Gao W, Xu L, et al. GbMPK3, a mitogen-activated protein kinase from cotton, enhances drought and oxidative stress tolerance in tobacco[J]. Plant Cell Tissue Organ Cult PCTOC, 2014, 116(2): 153-162. Wu J, Liang X Y, Lin M, et al. Comprehensive analysis of MAPK gene family in Populus trichocarpa and physiological characterization of PtMAPK3-1 in response to MeJA induction[J]. Physiol Plant, 2023, 175(1): e13869. Hann C T, Ramage S F, Negi H, et al. Dephosphorylation of the MAP kinases MPK6 and MPK3 fine-tunes responses to wounding and herbivory in Arabidopsis[J]. Plant Sci, 2024, 339: 111962. Zhou J G, Mu Q, Wang X Y, et al. Multilayered synergistic regulation of phytoalexin biosynthesis by ethylene, jasmonate, and MAPK signaling pathways in Arabidopsis[J]. Plant Cell, 2022, 34(8): 3066-3087. Qu R J, Wang S W, Wang X X, et al. The jasmonate-responsive SmMPK3-SmWRKY33 module positively regulates tanshinone biosynthesis in Salvia miltiorrhiza[J]. Plant Biotechnol J, 2026, 24(2): 384-401. Xu Y H, Zhang Z, Wang M X, et al. Identification of genes related to agarwood formation: Transcriptome analysis of healthy and wounded tissues of Aquilaria sinensis[J]. BMC Genomics, 2013, 14: 227. Sun P W, Xu Y H, Yu C C, et al. AsWRKY44 represses the wound-induced sesquiterpene biosynthetic gene ASS1 expression in Aquilaria sinensis (Lour.) Gilg[J]. J Exp Bot, 2019: erz469. Xu Y H, Lv F F, Sun P W, et al. Cloning and functional analysis of the promoterof the sesquiterpene synthase gene ASS1 in Aquilaria sinensis[J]. Biologia Plant, 2021, 65: 60-67. 刘娟, 韩晓敏, 梁良, 等. 濒危南药白木香悬浮细胞体系的建立[J]. 药学学报, 2014, 49(8): 1194-1199. Yoo S D, Cho Y H, Sheen J. Arabidopsis mesophyll protoplasts: A versatile cell system for transient gene expression analysis[J]. Nat Protoc, 2007, 2(7): 1565-1572. Zhang S, Klessig D F. MAPK cascades in plant defense signaling[J]. Trends Plant Sci, 2001, 6(11): 520-527. Bigeard J, Colcombet J, Hirt H. Signaling mechanisms in pattern-triggered immunity (PTI)[J]. Mol Plant, 2015, 8(4): 521-539. Frei dit Frey N, Garcia A V, Bigeard J, et al. Functional analysis of Arabidopsis immune-related MAPKs uncovers a role for MPK3 as negative regulator of inducible defences[J]. Genome Biol, 2014, 15(6): R87. Pitzschke A, Datta S, Persak H. Salt stress in Arabidopsis: Lipid transfer protein AZI1 and its control by mitogen-activated protein kinase MPK3[J]. Mol Plant, 2014, 7(4): 722-738. Melikant B, Giuliani C, Halbmayer-Watzina S, et al. The Arabidopsis thaliana MEK AtMKK6 activates the MAP kinase AtMPK13[J]. FEBS Lett, 2004, 576(1/2): 5-8. Zeng Q N, Sritubtim S, Ellis B E. AtMKK6 and AtMPK13 are required for lateral root formation in Arabidopsis[J]. Plant Signal Behav, 2011, 6(10): 1436-1439. Ligterink W, Kroj T, zur Nieden U, et al. Receptor-mediated activation of a MAP kinase in pathogen defense of plants[J]. Science, 1997, 276(5321): 2054-2057. Carrasco J L, Castelló M J, Naumann K, et al. Arabidopsis protein phosphatase DBP1 nucleates a protein network with a role in regulating plant defense[J]. PLoS One, 2014, 9(3): e90734. Wang J X, Ding H D, Zhang A Y, et al. A novel mitogen-activated protein kinase gene in maize (Zea mays), ZmMPK3, is involved in response to diverse environmental cues[J]. J Integr Plant Biol, 2010, 52(5): 442-452. Liang W W, Yang B, Yu B J, et al. Identification and analysis of MKK and MPK gene families in canola (Brassica napus L.)[J]. BMC Genomics, 2013, 14: 392. Liu J Z, Horstman H D, Braun E, et al. Soybean homologs of MPK4 negatively regulate defense responses and positively regulate growth and development[J]. Plant Physiol, 2011, 157(3): 1363-1378.)
ObjectiveAsMAPK1 and AsMAPK2 are two mitogen-activated protein kinase (MAPK) genes with unknown functions in Aquilaria sinensis. To investigate their fundamental characteristics and potential functions in wound-induced agarwood formation, in this study, we performed cloning, expression analysis, and preliminary in vitro functional assays on the two genes. Methods Using cDNA from subcultured A. sinensis callus as the template, the coding sequences of AsMAPK1 and AsMAPK2 were obtained by PCR amplification. Bioinformatics analysis was performed using relevant online tools and softwares. The prokaryotic expression vectors pGEX4T-1-AsMAPK1/2 were constructed, transformed into E. coli BL21 for expression, and induction conditions were optimized. The AsMAPK1-GFP and AsMAPK2-GFP fusion expression vectors were constructed and transformed into Arabidopsis thaliana protoplasts to observe their subcellular localization using confocal microscopy. The kinase activity of the purified proteins was detected using an in vitro phosphorylation assay. Results The AsMAPK1 (1 188 bp) and AsMAPK2 (1 128 bp) genes were successfully cloned. Bioinformatics analysis revealed that both encoded proteins contain typical kinase domains and belong to the MAPK family. Phylogenetic analysis indicated that AsMAPK1 is most closely related to the AtMAPK6 homolog in A. thaliana and belongs to Group A, while AsMAPK2 clusters with Group B AtMAPK13. SDS-PAGE detection showed successful induction of soluble recombinant proteins of approximately 71 000 and 69 000. Subcellular localization results demonstrated that both AsMAPK1-GFP and AsMAPK2-GFP are localized in the cytoplasm and nucleus. Further kinase activity analysis confirmed that both AsMAPK1 and AsMAPK2 undergo autophosphorylation and can phosphorylate the universal substrate MBP. Conclusion This study confirms that AsMAPK1 and AsMAPK2 are active protein kinases localized in both the cytoplasm and nucleus. The cloning, expression analysis, and kinase activity detection of the two AsMAPKs provide a foundation for further elucidating their roles in the signal transduction mechanism underlying wound-induced agarwood formation in A. sinensis.
YANG Shuwen, RONG Mei, WANG Siyu, SUN Yuxuan, XU Yanhong, WEI Jianhe.
Cloning, prokaryotic expression, and activity analysis of two MAPK genes in Aquilaria sinensis[J].
Chinese Traditional and Herbal Drugs,
2026
, 57
(6)
: 2265
-2277
.
DOI: 10.7501/j.issn.0253-2670.2026.06.021
李林海, 寿海洋, 马清温. 土沉香(瑞香科)的地理分布研究[J]. 安徽农业科学, 2012, 40(17): 9254-9256. 张争, 杨云, 魏建和, 等. 白木香结香机制研究进展及其防御反应诱导结香假说[J]. 中草药, 2010, 41(1): 156-159. Li W, Chen H Q, Wang H, et al. Natural products in agarwood and Aquilaria plants: Chemistry, biological activities and biosynthesis[J]. Nat Prod Rep, 2021, 38(3): 528-565. Wang S, Yu Z X, Wang C H, et al. Chemical constituents and pharmacological activity of agarwood and Aquilaria plants[J]. Molecules, 2018, 23(2): 342. 雷胄熙, 吴泽青, 林励, 等. 生物诱导法所产不同部位沉香挥发油成分分析[J]. 中药新药与临床药理, 2016, 27(1): 106-113. 林峰, 戴好富, 王辉, 等. 两批接菌法所产沉香挥发油化学成分的气相色谱-质谱联用分析[J]. 时珍国医国药, 2010, 21(8): 1901-1902. Liu Y Y, Chen H Q, Yang Y, et al. Whole-tree agarwood-inducing technique: An efficient novel technique for producing high-quality agarwood in cultivated Aquilaria sinensis trees[J]. Molecules, 2013, 18(3): 3086-3106. Jiang M, Zhang Y Z, Li P, et al. Mitogen-activated protein kinase and substrate identification in plant growth and development[J]. Int J Mol Sci, 2022, 23(5): 2744. Xu J, Zhang S Q. Mitogen-activated protein kinase cascades in signaling plant growth and development[J]. Trends Plant Sci, 2015, 20(1): 56-64. Zhang M M, Zhang S Q. Mitogen-activated protein kinase cascades in plant signaling[J]. J Integr Plant Biol, 2022, 64(2): 301-341. Mao G H, Meng X Z, Liu Y D, et al. Phosphorylation of a WRKY transcription factor by two pathogen-responsive MAPKs drives phytoalexin biosynthesis in Arabidopsis[J]. Plant Cell, 2011, 23(4): 1639-1653. Meng X Z, Xu J, He Y X, et al. Phosphorylation of an ERF transcription factor by Arabidopsis MPK3/MPK6 regulates plant defense gene induction and fungal resistance[J]. Plant Cell, 2013, 25(3): 1126-1142. Kishi-Kaboshi M, Seo S, Takahashi A, et al. The MAMP-responsive MYB transcription factors MYB30, MYB55 and MYB110 activate the HCAA synthesis pathway and enhance immunity in rice[J]. Plant Cell Physiol, 2018, 59(5): 903-915. Zhang D, Jiang S, Pan J, et al. The overexpression of a maize mitogen-activated protein kinase gene (ZmMPK5) confers salt stress tolerance and induces defence responses in tobacco[J]. Plant Biol, 2014, 16(3): 558-570. Long L, Gao W, Xu L, et al. GbMPK3, a mitogen-activated protein kinase from cotton, enhances drought and oxidative stress tolerance in tobacco[J]. Plant Cell Tissue Organ Cult PCTOC, 2014, 116(2): 153-162. Wu J, Liang X Y, Lin M, et al. Comprehensive analysis of MAPK gene family in Populus trichocarpa and physiological characterization of PtMAPK3-1 in response to MeJA induction[J]. Physiol Plant, 2023, 175(1): e13869. Hann C T, Ramage S F, Negi H, et al. Dephosphorylation of the MAP kinases MPK6 and MPK3 fine-tunes responses to wounding and herbivory in Arabidopsis[J]. Plant Sci, 2024, 339: 111962. Zhou J G, Mu Q, Wang X Y, et al. Multilayered synergistic regulation of phytoalexin biosynthesis by ethylene, jasmonate, and MAPK signaling pathways in Arabidopsis[J]. Plant Cell, 2022, 34(8): 3066-3087. Qu R J, Wang S W, Wang X X, et al. The jasmonate-responsive SmMPK3-SmWRKY33 module positively regulates tanshinone biosynthesis in Salvia miltiorrhiza[J]. Plant Biotechnol J, 2026, 24(2): 384-401. Xu Y H, Zhang Z, Wang M X, et al. Identification of genes related to agarwood formation: Transcriptome analysis of healthy and wounded tissues of Aquilaria sinensis[J]. BMC Genomics, 2013, 14: 227. Sun P W, Xu Y H, Yu C C, et al. AsWRKY44 represses the wound-induced sesquiterpene biosynthetic gene ASS1 expression in Aquilaria sinensis (Lour.) Gilg[J]. J Exp Bot, 2019: erz469. Xu Y H, Lv F F, Sun P W, et al. Cloning and functional analysis of the promoterof the sesquiterpene synthase gene ASS1 in Aquilaria sinensis[J]. Biologia Plant, 2021, 65: 60-67. 刘娟, 韩晓敏, 梁良, 等. 濒危南药白木香悬浮细胞体系的建立[J]. 药学学报, 2014, 49(8): 1194-1199. Yoo S D, Cho Y H, Sheen J. Arabidopsis mesophyll protoplasts: A versatile cell system for transient gene expression analysis[J]. Nat Protoc, 2007, 2(7): 1565-1572. Zhang S, Klessig D F. MAPK cascades in plant defense signaling[J]. Trends Plant Sci, 2001, 6(11): 520-527. Bigeard J, Colcombet J, Hirt H. Signaling mechanisms in pattern-triggered immunity (PTI)[J]. Mol Plant, 2015, 8(4): 521-539. Frei dit Frey N, Garcia A V, Bigeard J, et al. Functional analysis of Arabidopsis immune-related MAPKs uncovers a role for MPK3 as negative regulator of inducible defences[J]. Genome Biol, 2014, 15(6): R87. Pitzschke A, Datta S, Persak H. Salt stress in Arabidopsis: Lipid transfer protein AZI1 and its control by mitogen-activated protein kinase MPK3[J]. Mol Plant, 2014, 7(4): 722-738. Melikant B, Giuliani C, Halbmayer-Watzina S, et al. The Arabidopsis thaliana MEK AtMKK6 activates the MAP kinase AtMPK13[J]. FEBS Lett, 2004, 576(1/2): 5-8. Zeng Q N, Sritubtim S, Ellis B E. AtMKK6 and AtMPK13 are required for lateral root formation in Arabidopsis[J]. Plant Signal Behav, 2011, 6(10): 1436-1439. Ligterink W, Kroj T, zur Nieden U, et al. Receptor-mediated activation of a MAP kinase in pathogen defense of plants[J]. Science, 1997, 276(5321): 2054-2057. Carrasco J L, Castelló M J, Naumann K, et al. Arabidopsis protein phosphatase DBP1 nucleates a protein network with a role in regulating plant defense[J]. PLoS One, 2014, 9(3): e90734. Wang J X, Ding H D, Zhang A Y, et al. A novel mitogen-activated protein kinase gene in maize (Zea mays), ZmMPK3, is involved in response to diverse environmental cues[J]. J Integr Plant Biol, 2010, 52(5): 442-452. Liang W W, Yang B, Yu B J, et al. Identification and analysis of MKK and MPK gene families in canola (Brassica napus L.)[J]. BMC Genomics, 2013, 14: 392. Liu J Z, Horstman H D, Braun E, et al. Soybean homologs of MPK4 negatively regulate defense responses and positively regulate growth and development[J]. Plant Physiol, 2011, 157(3): 1363-1378.
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