Article(id=1321785499325420413, tenantId=1146029695717560320, journalId=1316781310003478668, issueId=1321785454333121105, articleNumber=null, orderNo=null, doi=10.11983/CBB24019, pmid=null, cstr=32102.14.CBB24019, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=research-article, receivedDate=1707062400000, receivedDateStr=2024-02-05, revisedDate=null, revisedDateStr=null, acceptedDate=1724083200000, acceptedDateStr=2024-08-20, onlineDate=1793067820626, onlineDateStr=2026-10-27, pubDate=1736438400000, pubDateStr=2025-01-10, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1793067820626, onlineIssueDateStr=2026-10-27, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1793067820626, creator=13701087609, updateTime=1793067820626, updator=13701087609, issue=Issue{id=1321785454333121105, tenantId=1146029695717560320, journalId=1316781310003478668, year='2025', volume='60', issue='1', pageStart='1', pageEnd='143', issueExtLink='null', onlineDate='null', pubDate='1736438400000', pubDateStr='2025-01-10', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=null, createTime=1793067809899, creator='13701087609', updateTime=1793068026855, updator='13701087609', preIssue=null, nextIssue=null, articleTotal=null, ext={EN=IssueExt(id=1321786364417397082, tenantId=1146029695717560320, journalId=1316781310003478668, issueId=1321785454333121105, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1321786364417397083, tenantId=1146029695717560320, journalId=1316781310003478668, issueId=1321785454333121105, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null, downloadFileDto=null}, startPage=49, endPage=61, ext={EN=ArticleExt(id=1321785499530941310, articleId=1321785499325420413, tenantId=1146029695717560320, journalId=1316781310003478668, language=EN, title=Analysis of Expression Characteristics and Identification of Interaction Proteins of BnaABF2 Transcription Factor in Brassica napus, columnId=1321785481268933599, journalTitle=Chinese Bulletin of Botany, columnName=EXPERIMENTAL COMMUNICATIONS, runingTitle=null, highlight=null, articleAbstract=

ABF transcription factors are collectively referred to as basic leucine zipper proteins that can specifically recognize and bind to ABA-responsive elements (ABRE), participating in ABA signal transduction and serving as regulators of ABA signal transcriptional responses. This study analyzed the protein encoded by the BnaABF2 gene in Brassica napus. Subcellular localization results showed that the BnaABF2 protein is localized in the nucleus. Analysis of transcriptional activity in the yeast system indicated that BnaABF2 has no transcriptional activation activity; qRT-PCR detection revealed that the expression level of BnaABF2 is highest in leaves. We also found that ABA treatment, simulated drought, and salt stress can induce the expression of BnaABF2; BiFC results showed that BnaMPK1/2/6/7/9/12/13 can interact with BnaABF2. Dual-LUC results suggested that BnaMPK7 may enhance the transcriptional regulation of BnaABF2 on downstream target genes through phosphorylation. This study initially explored the basic characteristics and interacting proteins of the transcription factor BnaABF2, providing theoretical guidance for understanding its functions and mechanisms.

, authors=Liuqing Yang, Jin Wang, Jingli Yan, Qinqin Chen, Haokun Cheng, Chun Li, Peiyu Zhao, Bo Yang, Yuanqing Jiang*, authorsList=Liuqing Yang, Jin Wang, Jingli Yan, Qinqin Chen, Haokun Cheng, Chun Li, Peiyu Zhao, Bo Yang, Yuanqing Jiang, authorCompany=null, correspAuthors=Yuanqing Jiang, authorNote=

†These authors contributed equally to this paper

, correspAuthorsNote=
* E-mail:
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ABF转录因子是能够特异识别并结合ABA响应元件(ABRE)的碱性亮氨酸拉链蛋白的统称, 参与ABA信号转导。通过对甘蓝型油菜(Brassica napus) BnaABF2基因编码蛋白进行分析, 亚细胞定位结果显示, BnaABF2蛋白定位于细胞核; 酵母系统转录活性分析表明, BnaABF2无转录激活活性; qRT-PCR检测发现, BnaABF2在叶中的表达量最高。此外, 还发现ABA处理、模拟干旱和盐胁迫能够诱导BnaABF2的表达; BiFC结果显示, BnaMPK1/2/6/7/9/12/13能与BnaABF2相互作用。Dual-LUC结果表明, BnaMPK7可能通过磷酸化增强BnaABF2对下游靶基因的转录调控。该研究初步探索了转录因子BnaABF2的基本特性与互作蛋白, 对理解其功能与机制具有一定的理论价值。

, authors=杨柳卿†, 王劲†, 燕敬利, 陈芹芹, 程浩坤, 李春, 赵培玉, 杨博, 江元清*, authorsList=杨柳卿, 王劲, 燕敬利, 陈芹芹, 程浩坤, 李春, 赵培玉, 杨博, 江元清, authorCompany=null, correspAuthors=江元清, authorNote=

†共同第一作者。

, correspAuthorsNote=
* 江元清, 男, 汉族, 湖南省桃源县人, 教授, 博士生导师。1998年6月本科毕业于湖南农业大学资环学院; 2001年6月硕士研究生毕业于中国农业大学生物学院生物化学与分子生物学专业。2001.07-2003.08在北京市农林科学院农业生物技术研究中心工作(助理研究员), 从事扁桃基因功能分析相关研究。2003.09-2007.11在加拿大阿尔伯特大学(University of Alberta)生物科学系学习(植物生物学专业), 获博士学位。2007.12-2010.11先后在阿尔伯特大学生物科学系和美国加州大学伯克利分校(University of California-Berkeley)植物与微生物学系做博士后研究。2010年12月就职于西北农林科技大学生命科学学院, 从事教学与科研工作。E-mail:
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Abiotic stress signaling and responses in plants. Cell 167, 313-324., articleTitle=Abiotic stress signaling and responses in plants, refAbstract=null)], funds=[Fund(id=1321785509991535578, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, awardId=32171953, language=CN, fundingSource=国家自然科学基金(32171953), fundOrder=null, country=null)], companyList=[AuthorCompany(id=1321785503276454800, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, xref=null, ext=[AuthorCompanyExt(id=1321785503284843409, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, companyId=1321785503276454800, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=College of Life Sciences, Northwest A & F University, Yangling 712100, China), AuthorCompanyExt(id=1321785503293232018, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, companyId=1321785503276454800, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=西北农林科技大学生命科学学院, 杨凌 712100)])], figs=[ArticleFig(id=1321785508167013322, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, language=EN, label=Figure 1, caption=Cis-elements of two BnaABF2 promoters

ABRE: Abscisic acid responsiveness element; ARE: Anaerobic induction element; Light: Light responsiveness element; TCA- element: Salicylic acid responsiveness element; TC-rich repeats: Defense and stress responsiveness element; TGA-element: Auxin responsiveness element; CGTCA-motif: MeJA-responsiveness motif; TGACG-motif: Jasmonic acid responsiveness motif; Circadian: Circadian rhythm

, figureFileSmall=zH2mLJUS/9jdYZ0i/FToKw==, figureFileBig=rXYzbs20zHMGIREYVYpvAg==, tableContent=null), ArticleFig(id=1321785508305425355, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, language=CN, label=图1, caption=2个BnaABF2基因启动子的顺式作用元件

ABRE: 脱落酸响应元件; ARE: 厌氧诱导元件; Light: 光响应元件; TCA-element: 水杨酸响应元件; TC-rich repeats: 防御和应激反应元件; TGA-element: 生长素响应元件; CGTCA-motif: 茉莉酸甲酯响应基序; TGACG-motif: 茉莉酸响应基序; Circadian: 昼夜节律

, figureFileSmall=zH2mLJUS/9jdYZ0i/FToKw==, figureFileBig=rXYzbs20zHMGIREYVYpvAg==, tableContent=null), ArticleFig(id=1321785508578055116, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, language=EN, label=Figure 2, caption=Sequence alignment of conserved domains of BnaABF2 with ABF from other species

Left: Protein conserved motifs of ABF2/3 alignment; Middle: ABF2/3 protein conserved domain alignment; Right: Transcript structure alignment of coding region (CDS) and untranslated region (UTR) of ABF2/3.

, figureFileSmall=XrvuIDrYAC9SSxc2cij6FA==, figureFileBig=fq9lhM/Mx6ONIYxJY8mGOA==, tableContent=null), ArticleFig(id=1321785508657746893, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, language=CN, label=图2, caption=BnaABF2/3与其它物种ABF保守结构域序列比对

左列: ABF2/3蛋白保守基序比对; 中列: ABF2/3蛋白保守结构域比对; 右列: ABF2/3编码区(coding sequence, CDS)与非翻译区(untranslated region, UTR)的转录本结构比对。

, figureFileSmall=XrvuIDrYAC9SSxc2cij6FA==, figureFileBig=fq9lhM/Mx6ONIYxJY8mGOA==, tableContent=null), ArticleFig(id=1321785508733244366, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, language=EN, label=Figure 3, caption=A phylogenetic tree of BnaABF2 and ABF proteins from other species

The branches of the evolutionary tree showed genetic distance with a confidence level of 100.

, figureFileSmall=iE7Py1jhFD5kC+y8i7mSkw==, figureFileBig=d8EkQcRjqc2bA3DwjcAnpw==, tableContent=null), ArticleFig(id=1321785508804547535, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, language=CN, label=图3, caption=BnaABF2与其它物种ABF蛋白的系统进化树

进化树分支显示遗传距离, 置信度为100。

, figureFileSmall=iE7Py1jhFD5kC+y8i7mSkw==, figureFileBig=d8EkQcRjqc2bA3DwjcAnpw==, tableContent=null), ArticleFig(id=1321785508871656400, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, language=EN, label=Figure 4, caption=Subcellular localization of BnaABF2

pYJHF-BnaABF2 or pYJHF (control) plasmid were infiltrated into tobacco leaves, and fluorescence signals were observed two days later. GFP: Green fluorescent protein signal; NLS-mCherry: Nuclear localization marker; Bright: Bright field; Merge: Superimposed field. Bars=50 μm

, figureFileSmall=p5WQ//lQgNJcvCsW1rjOSg==, figureFileBig=bxCj/IrlzDFjrz1sjKb1bA==, tableContent=null), ArticleFig(id=1321785508947153873, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, language=CN, label=图4, caption=BnaABF2的亚细胞定位

将重组质粒pYJHF-BnaABF2或对照质粒pYJHF转染烟草叶片, 2天后观察荧光信号。GFP: 绿色荧光蛋白信号; NLS-mCherry: 核定位标记; Bright: 明场; Merge: 叠加场。Bars=50 μm

, figureFileSmall=p5WQ//lQgNJcvCsW1rjOSg==, figureFileBig=bxCj/IrlzDFjrz1sjKb1bA==, tableContent=null), ArticleFig(id=1321785509031039954, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, language=EN, label=Figure 5, caption=Analysis of transcriptional activity of BnaABF2 in yeast system

BD (negative control), BD-BnaABF2, BD-BnaMYB111 (positive control), BD-BnaABF2 5m, BD-BnaABF2 5D and BD-BnaABF2 Co were transferred into yeast for titration experiment. The dilution gradients of yeast cells were marked with black triangles.

, figureFileSmall=/RYwulj/gKk9VCOJX5FwlQ==, figureFileBig=hVWFaNvuXC/OL97YNrpn0Q==, tableContent=null), ArticleFig(id=1321785509127508947, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, language=CN, label=图5, caption=BnaABF2的酵母系统转录活性分析

分别将BD (阴性对照)、BD-BnaABF2、BD-BnaMYB111 (阳性对照)、BD-BnaABF2 5m、BD-BnaABF2 5D和BD-BnaABF2 Co转入酵母进行酵母滴定实验。酵母细胞稀释梯度使用黑色三角形标示。

, figureFileSmall=/RYwulj/gKk9VCOJX5FwlQ==, figureFileBig=hVWFaNvuXC/OL97YNrpn0Q==, tableContent=null), ArticleFig(id=1321785509211395028, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, language=EN, label=Figure 6, caption=Expression patterns of BnaABF2 gene in different tissues and under different stresses and plant growth regulator treatments

(A) The expression of BnaABF2 in different tissues; (B) The expression of BnaABF2 under various stresses and plant growth regulator treatments, the treatments included 50 μmol·L-1 ABA, 15% PEG8000, cold (4°C), heat (38°C) and salinity (200 mmol·L-1 NaCl). Values are means±SE. The significant differences in t-test were indicated by asterisk, * P<0.05; ** P<0.01

, figureFileSmall=gcqPMlZHH4BCA9qr8Bvk4Q==, figureFileBig=uXoYOnjOBpDDRaWFvx/zcg==, tableContent=null), ArticleFig(id=1321785509303669717, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, language=CN, label=图6, caption=BnaABF2在不同组织及多种逆境与植物生长调节剂处理下的表达分析

(A) BnaABF2在不同组织中的表达情况; (B) BnaABF2在多种逆境与植物生长调节剂处理下的表达情况, 处理条件为脱落酸(abscisic acid, ABA) (50 μmol·L-1)、模拟干旱(15% PEG8000)、冷害(4°C)、热害(38°C)和盐害(200 mmol·L-1 NaCl)。数据为平均值±标准误。星号表示t检验存在显著性差异, * P<0.05; ** P<0.01

, figureFileSmall=gcqPMlZHH4BCA9qr8Bvk4Q==, figureFileBig=uXoYOnjOBpDDRaWFvx/zcg==, tableContent=null), ArticleFig(id=1321785509584688086, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, language=EN, label=Figure 7, caption=Screening of interaction between BnaABF2 and BnaMPK proteins through bimolecular fluorescence complementation assay

The plasmids of BnaABF2-YFPc and YFPn-BnaMPK1/2/6/ 7/9/12/13 and NLS-mCherry were co-transfected into tobacco (28 d). Four days later, the fluorescence signal was observed. YFPn: Yellow fluorescent protein N-terminal sequence; YFPc: Yellow fluorescent protein C-terminal sequence; YFP: Yellow fluorescent protein signal; NLS-mCherry: Nuclear localization red fluorescence signal; Bright: Bright field; Merge: Superimposed field. Bars=50 μm

, figureFileSmall=yerD6DDb9q6l3sGyuOcsCQ==, figureFileBig=EjfsnEXmT1hlPVeZAP7E7A==, tableContent=null), ArticleFig(id=1321785509718905815, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, language=CN, label=图7, caption=双分子荧光互补实验检测油菜的BnaABF2蛋白与BnaMPKs蛋白间相互作用

将含有BnaABF2-YFPc和YFPn-BnaMPK1/2/6/7/9/12/13的质粒与NLS-mCherry共同转染28天本氏烟草, 4天后观察荧光信号。YFPn: 黄色荧光蛋白N端序列; YFPc: 黄色荧光蛋白C端序列; YFP: 黄色荧光蛋白信号; NLS-mCherry: 核定位红色荧光信号; Bright: 明场; Merge: 叠加场。Bars=50 μm

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(A) A schematic diagram of the reporter and effector; (B) The LUC/REN ratio is used to indicate the ability of the effector to activate the reporter (dpi: Days post-infiltration). Different lowercase letters indicate significant differences between different treatments (P<0.05).

, figureFileSmall=90BKnnysSEnS7VaffpN94A==, figureFileBig=yikJA4vRAIC4ZJKyFpn+3A==, tableContent=null), ArticleFig(id=1321785509878289369, tenantId=1146029695717560320, journalId=1316781310003478668, articleId=1321785499325420413, language=CN, label=图8, caption=双荧光素酶报告系统检测脱落酸(ABA)处理下BnaABF2对AtRD29B启动子转录活性的影响

(A) 报告子和效应子示意图; (B) LUC/REN比值表示效应子激活报告子的能力, 数值越大, 能力越强(dpi: 注射后的天数)。不同小写字母表示各处理间差异显著(P<0.05)。

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甘蓝型油菜转录因子BnaABF2的表征分析及互作蛋白鉴定
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杨柳卿 † , 王劲 † , 燕敬利 , 陈芹芹 , 程浩坤 , 李春 , 赵培玉 , 杨博 , 江元清 *
植物学报 | 研究报告 2025,60(1): 49-61
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植物学报 |研究报告 2025 , 60 (1) : 49 -61
甘蓝型油菜转录因子BnaABF2的表征分析及互作蛋白鉴定
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杨柳卿†, 王劲†, 燕敬利, 陈芹芹, 程浩坤, 李春, 赵培玉, 杨博, 江元清*
作者信息
  • 西北农林科技大学生命科学学院, 杨凌 712100
通讯作者:
* 江元清, 男, 汉族, 湖南省桃源县人, 教授, 博士生导师。1998年6月本科毕业于湖南农业大学资环学院; 2001年6月硕士研究生毕业于中国农业大学生物学院生物化学与分子生物学专业。2001.07-2003.08在北京市农林科学院农业生物技术研究中心工作(助理研究员), 从事扁桃基因功能分析相关研究。2003.09-2007.11在加拿大阿尔伯特大学(University of Alberta)生物科学系学习(植物生物学专业), 获博士学位。2007.12-2010.11先后在阿尔伯特大学生物科学系和美国加州大学伯克利分校(University of California-Berkeley)植物与微生物学系做博士后研究。2010年12月就职于西北农林科技大学生命科学学院, 从事教学与科研工作。E-mail:
作者简介:

†共同第一作者。

Analysis of Expression Characteristics and Identification of Interaction Proteins of BnaABF2 Transcription Factor in Brassica napus
Liuqing Yang, Jin Wang, Jingli Yan, Qinqin Chen, Haokun Cheng, Chun Li, Peiyu Zhao, Bo Yang, Yuanqing Jiang*
Affiliations
  • College of Life Sciences, Northwest A & F University, Yangling 712100, China
  • Corresponding Author:
出版时间: 2025-01-10 doi: 10.11983/CBB24019
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ABF转录因子是能够特异识别并结合ABA响应元件(ABRE)的碱性亮氨酸拉链蛋白的统称, 参与ABA信号转导。通过对甘蓝型油菜(Brassica napus) BnaABF2基因编码蛋白进行分析, 亚细胞定位结果显示, BnaABF2蛋白定位于细胞核; 酵母系统转录活性分析表明, BnaABF2无转录激活活性; qRT-PCR检测发现, BnaABF2在叶中的表达量最高。此外, 还发现ABA处理、模拟干旱和盐胁迫能够诱导BnaABF2的表达; BiFC结果显示, BnaMPK1/2/6/7/9/12/13能与BnaABF2相互作用。Dual-LUC结果表明, BnaMPK7可能通过磷酸化增强BnaABF2对下游靶基因的转录调控。该研究初步探索了转录因子BnaABF2的基本特性与互作蛋白, 对理解其功能与机制具有一定的理论价值。

甘蓝型油菜  /  ABF2  /  亚细胞定位  /  MPK  /  互作蛋白

ABF transcription factors are collectively referred to as basic leucine zipper proteins that can specifically recognize and bind to ABA-responsive elements (ABRE), participating in ABA signal transduction and serving as regulators of ABA signal transcriptional responses. This study analyzed the protein encoded by the BnaABF2 gene in Brassica napus. Subcellular localization results showed that the BnaABF2 protein is localized in the nucleus. Analysis of transcriptional activity in the yeast system indicated that BnaABF2 has no transcriptional activation activity; qRT-PCR detection revealed that the expression level of BnaABF2 is highest in leaves. We also found that ABA treatment, simulated drought, and salt stress can induce the expression of BnaABF2; BiFC results showed that BnaMPK1/2/6/7/9/12/13 can interact with BnaABF2. Dual-LUC results suggested that BnaMPK7 may enhance the transcriptional regulation of BnaABF2 on downstream target genes through phosphorylation. This study initially explored the basic characteristics and interacting proteins of the transcription factor BnaABF2, providing theoretical guidance for understanding its functions and mechanisms.

Brassica napus  /  ABF2  /  subcellular localization  /  MPK  /  interaction proteins
杨柳卿, 王劲, 燕敬利, 陈芹芹, 程浩坤, 李春, 赵培玉, 杨博, 江元清. 甘蓝型油菜转录因子BnaABF2的表征分析及互作蛋白鉴定. 植物学报, 2025 , 60 (1) : 49 -61 . DOI: 10.11983/CBB24019
Liuqing Yang, Jin Wang, Jingli Yan, Qinqin Chen, Haokun Cheng, Chun Li, Peiyu Zhao, Bo Yang, Yuanqing Jiang. Analysis of Expression Characteristics and Identification of Interaction Proteins of BnaABF2 Transcription Factor in Brassica napus[J]. Chinese Bulletin of Botany, 2025 , 60 (1) : 49 -61 . DOI: 10.11983/CBB24019
甘蓝型油菜(Brassica napus)隶属十字花科芸薹属, 是我国主要油料作物之一, 主要种植于长江中下游地区。其祖先由野甘蓝(B. oleracea)和白菜(B. rapa)杂交, 而后染色体加倍形成异源四倍体作物(Chalhoub et al., 2014)。该型油菜具有含油量高和产量高等优点, 但耐寒性和抗旱性差且不耐瘠薄。因此, 深入探究甘蓝型油菜的抗逆基因及其响应机制, 对于增强其抵御非生物胁迫的能力、提升产量和品质及指导育种工作均具有重要意义。
脱落酸(abscisic acid, ABA)是一种调节植物生长发育和胁迫反应的重要植物激素, 调控种子发育和萌发等一系列生理过程。脱落酸信号转导途径是植物在干旱以及盐胁迫下生存和适应的核心机制(Zhu, 2016)。研究人员对ABA诱导基因的启动子区域进行序列分析, 发现1个保守的顺式作用元件, 称为ABA反应元件(ABA-responsive element, ABRE)。与ABRE结合的转录因子, 称为ABA反应元件结合因子(ABA-responsive element binding factors, AREBs/ ABFs)。ABFs/AREBs属于bZIP转录因子家族, 参与ABA信号转导途径并作为ABA信号转录应答的调节因子。ABA信号调控在植物逆境适应中占核心地位, 对植物的生存以及生长具有重要意义(周玉萍等, 2022)。拟南芥(Arabidopsis thaliana)的ABFs家族成员主要在营养组织中表达, 并且表达受到各种非生物胁迫诱导(Choi et al., 2000; Uno et al., 2000)。此外, 它们有不同的应激诱导模式, 表明不同的AtABF可能在不同的应激反应途径中发挥作用; 即AtABF1在冷信号通路中, AtABF2和AtABF3在盐信号通路中, AtABF4在冷、盐和干旱信号通路中行使功能(Kim et al., 2004; Yoshida et al., 2010)。已有研究表明, AtABF2和AtABF4的最大转录活性依赖ABA的翻译后修饰(可能是磷酸化修饰) (Uno et al., 2000)。
丝裂原活化蛋白激酶(mitogen-activated protein kinase, MAPK)级联通路是植物中研究最多的信号机制之一, 在联系信号感知与多种细胞和适应性反应中起重要作用。MAPK模块通常至少包含3种蛋白激酶(MAP3K、MKK和MAPK (MPK))的组合, 其以顺序磷酸化依赖性方式被激活(Colcombet and Hirt, 2008)。激活的MAPK包括磷酸化的多种下游成分(如转录因子和乙烯合成酶), 以此调节它们的活性、稳定性或定位(Rodriguez et al., 2010)。其中, 拟南芥的MPK3、MPK4和MPK6是研究最多的3个MPK基因(Xu and Zhang, 2015)。
MPKs可磷酸化包括多个不同家族转录因子在内的多种底物(Wang et al., 2007; Mao et al., 2011)。例如, AtMPK3和AtMPK6磷酸化AtERF72的151位丝氨酸位点, 提高其转录调控活性以及植株的植保素含量和对灰霉菌(Botrytis cinerea)的抗性(Li et al., 2022)。AtMYB75是AtMPK4的底物, 被MPK4磷酸化后会导致光诱导的花青素积累(Li et al., 2016)。冷信号的转录抑制因子AtMYB15的活性受到AtMPK6介导的磷酸化调节(Kim et al., 2017)。本氏烟草(Nicotiana benthamiana) MPK对WRKYs的磷酸化促进WRKY依赖的RBOHB表达, 导致活性氧(reactive oxygen species, ROS)爆发(Adachi et al., 2015)。非生物胁迫能诱导AREB/ABF基因表达, 且有研究表明ABF2也受到ABA依赖的磷酸化调控, 需要ABA才能完全激活, 且其活性受到ABA依赖的保守结构域的多位点磷酸化调节(Fujita et al., 2005; Furihata et al., 2006)。作为脯氨酸导向的激酶, 激活的MAPK一般在脯氨酸之前的丝氨酸或苏氨酸残基上磷酸化其底物(Zhang and Zhang, 2022)。本实验室前期克隆了甘蓝型油菜中的12个BnaMPKs、7个BnaMKKs以及66个Bna-MPKKK, 并对其表达模式以及上下游互作蛋白等进行了初步探索(Liang et al., 2013; Sun et al., 2014), 为进一步研究其功能提供了思路。
基于本实验室前期的转录组测序结果, 我们聚焦于甘蓝型油菜BnaABFs家族, 筛选并克隆了表达差异显著的BnaABF2基因。通过生物信息学方法, 对BnaABF2基因的序列特征和系统进化关系进行分析。构建重组质粒并通过农杆菌介导技术, 在本氏烟草中实现了基因的瞬时表达, 进而观察基因在亚细胞水平的定位情况。同时, 我们利用实时荧光定量PCR技术检测了BnaABF2在不同组织器官的表达特点以及在不同非生物胁迫下的表达模式; 初步探索了BnaMPKs与BnaABF2的互作关系。该研究揭示了逆境胁迫下甘蓝型油菜BnaABF2基因的响应机制, 为进一步探索其抗逆功能奠定了基础。
本研究所用烟草为本氏烟草(Nicotiana benthamiana L.), 所用甘蓝型油菜(Brassica napus L.)为双单倍体DH12075; 温室光周期为14小时光照/10小时黑暗, 湿度介于60%-70%之间, 温度为22°C, 光照强度约为150 μmol·m-2·s-1。
本研究使用的基因组数据与蛋白序列文件分别来自甘蓝型油菜多组学信息资源数据库(https://yanglab.hzau.edu.cn/BnIR)和NCBI数据库(https://www.ncbi.nlm.nih.gov)。基于油菜品种Westar的蛋白组数据对ABF2测序结果进行本地BLAST, 随后利用BioLinux系统的检索功能提取ABF2同源基因上游1 500 bp的启动子序列, 并使用PlantCARE在线工具(https://bioin-formatics.psb.ugent.be/webtools/plantcare/html)进行预测, 以确定所有启动子序列范围内的顺式作用元件, 采用TBtools-II v2.034软件将ABF2同源基因启动子的顺式元件预测结果可视化。
从NCBI数据库下载拟南芥、水稻(Oryza sativa)和玉米(Zea mays)中ABF2同源基因的蛋白序列, 将油菜的蛋白序列与上述作物的同源序列合并进行系统进化分析。使用MEGA 7.0软件中的Clustal W模块对所有蛋白序列进行多序列联配, 通过最大似然估计(maximum likelihood estimation, MLE)构建系统进化树, 并设置“Possion model”作为Model/ Method参数。为处理缺失数据(missing data), 将缺失数据处理的参数设置为“Partial deletion”。为提高结果的稳定性, 进行了1 000次的Bootstrap校验。最后, 利用iTOL在线工具对系统进化分析结果可视化。
根据数据库中的gff基因注释文件, 得到上述作物ABF2同源基因的转录本信息; 基于上述作物的蛋白组序列信息, 利用BioLinux系统的MEME软件进行motif分析, 得到整体蛋白基序数据(nmotif参数设置为10, minw参数设置为6, maxw参数设置为50); 利用NCBI网站的CD-search在线工具对上述基因的蛋白序列进行结构域分析, 成功获得蛋白保守结构域信息。最后, 利用TBtools软件中的Gene Structure View模块进行可视化整合。
将BnaABF2基因的编码区序列构建至融合绿色荧光蛋白报告基因的表达载体pYJHF上, 所用引物为BnaABF2-BamHI-F: 5′-cgggatccATGAAATGCATGA TTTTAGTG-3′, BnaABF2-SalI-R: 5′-cgcgtcgacCC AAGGTCCTGACTCTGTCCT-3′。通过冻融法将构建好的重组质粒和p19 (来自番茄丛矮病毒)导入农杆菌(Agrobacterium tumefaciens) GV3101感受态细胞, 挑取阳性单菌落进行过夜培养。过夜培养物用含乙酰丁香酮的缓冲液(10 mmol·L-1 MES-KOH (pH5.6), 10 mmol·L-1 MgCl2, 150 mmol·L-1乙酰丁香酮)重悬至OD600=1; 黑暗条件下静置2-4小时后, 注射28天烟草叶片; 2天后使用激光共聚焦显微镜(Leica, 德国徕卡)观察(张翰风, 2019)。
构建重组质粒BD (pGBKT7)-BnaABF2、BD-BnaABF2 5m、BD-BnaABF2 5D和BD-BnaABF2 Co (所用引物序列同1.3节, 仅将正向引物酶切位点改为EcoRI), 并将其转入酵母感受态细胞AH109。然后, 检测转化后的酵母菌在营养缺陷型培养基上的生长情况。
营养缺陷型培养基包括一缺培养基(缺少色氨酸的SD-W)、二缺培养基(同时缺少色氨酸和组氨酸的SD-WH, 并在SD-WH基础上添加了3-AT (3-amino- 1,2,4-triazole, Sigma-Aldrich)的SD-WH+3-AT)和三缺培养基(同时缺少色氨酸、组氨酸和腺嘌呤半硫酸盐的SD-WHA)。其中, 3-AT是一种抑制剂, 工作浓度为2.5和5 mmol·L-1。
通过PEG4000-LiAc法, 将重组质粒BD-BnaABF2、BD-BnaABF2 5m、BD-BnaABF2 5D和BD- BnaABF2 Co与BD-BnaMYB111 (阳性对照)及空载体BD (阴性对照)分别导入酵母感受态细胞AH109。将转化后的酵母菌涂布于SD-W培养基上, 并在30°C下培养2天。待菌落长至合适大小后, 挑取单菌落接种于YEPD液体培养基, 30°C下, 于摇床上以200 r·min-1的速度继续培养, 直至OD600>1.5。使用分光光度计测定OD600, 以确定培养物的浓度。对培养物进行浓度梯度稀释, 并将稀释后的培养物滴定至不同类型的营养缺陷型培养基(SD-W、SD-WH、SD- WH+3-AT和SD-WHA)。30°C下培养并拍照。
将待测靶基因BnaMPKs和BnaABF2的编码区分别构建至35S-SPYNE(R)173和35S-SCYCE中(BnaMPKs所用引物参照Liang等(2013)所述, BnaABF2所用引物序列同1.3节, 仅将正向引物酶切位点改为SpeI), 并转入农杆菌GV3101, 挑取阳性菌落于YEB液体培养基中, 28°C振荡培养16小时。用含乙酰丁香酮的缓冲液重悬菌体至OD600=1, 将YFPN、YFPC、P19和NLS-mCherry (核定位marker)的重悬液按照体积比为1:1:1:1 (v/v/v/v)的比例混合, 黑暗下静置2-4小时后, 注射21天的本氏烟草叶片, 3天后使用激光共聚焦显微镜观察(Leica TCS SP8, 德国徕卡), 在激发光波长488 nm下观察黄色荧光蛋白YFP信号的有无和强弱(Niu et al., 2016)。
根据BnaABF2的cDNA序列, 设计定量PCR引物, BnaABF2-qPCR-F: 5′-AGGCGTTGTTGGTGCTGT- GTCTCC-3′, BnaABF2-qPCR-R: 5′-CCTTGCGGCT- GATTCTCGGTTCT-3′。以不同组织部位或逆境处理后的油菜cDNA为模板, 以BnaUBC9和BnaUP1为内参基因(引物序列见Liang等(2013)文献所述)进行qRT-PCR反应。反应程序为: 95°C3分钟; 95°C10秒, 60°C30秒, 40个循环。组织特异性表达的检测包括根、茎、叶、花蕾、花和果荚6种组织/器官; 非生物逆境处理包括50 μmol·L-1 ABA、模拟干旱(15% PEG8000)、盐害(200 mmol·L-1 NaCl)、热害(38°C)和冷害(4°C)。每个处理设4次生物学重复。
设计突变引物, 使用BnaABF2基因的正向引物F和反向突变型引物Rm进行Pfu高保真酶介导的PCR反应, 获得引入突变位点的短片段; 使用BnaABF2基因的反向引物R和Fm进行PCR反应, 获得另一条引入突变位点的短片段, 然后进行胶回收。通过这种方式, 可分别获得2条含有突变位点的短片段。将获得的2条短片段混合, 按循环参数1 (94°C3分钟; 94°C30秒, 60°C1分钟, 72°C每kb 1分钟, 8个循环; 72°C10分钟)进行PCR反应。步骤1的主要目的是促进2条短片段在突变位点的重叠序列互补配对, 从而形成杂交双链结构。随后, 加入基因两端的正反向引物(F和R), 按循环参数2 (94°C3分钟; 94°C30秒, 50°C1分钟, 72°C每kb 1分钟, 35个循环; 72°C10分钟)进行PCR反应。步骤2的主要目的是在杂交双链基础上, 使用BnaABF2基因的正向和反向引物进行PCR扩增, 从而获得带有突变位点的完整基因片段, 然后进行克隆与测序验证。
将目的基因BnaABF2及其突变序列BnaABF2 5m克隆到载体pBGHF上作为效应子, 将BnaMPK7克隆到载体pYJHA上。将从gDNA扩增获得的启动子ProAtRD29B克隆到载体pGreenII 0800-LUC上作为报告子。以报告子上35S启动子驱动的海肾荧光素酶基因为内参, 将重组质粒分别导入GV3101感受态细胞, 挑取阳性克隆并在YEB液体培养基中培养, 28°C条件下振荡培养16小时, 使菌落充分生长。用含乙酰丁香酮的缓冲液重悬菌体至OD600=0.5, 按照BnaMPK7: aABF2 (BnaABF2 5m):oAtRD29B=3:6:1 (v/v/v)混合均匀, 对照为NLS-GFP:BnaABF2 (或者BnaABF2 5m):ProAtRD29B=3:6:1 (v/v/v)。黑暗下静置2-4小时后注射28天本氏烟草叶片。处理2天后收取样品, 使用双荧光素酶报告分析(dualluciferase reporter system) (Promega)试剂和GloMax 20/20 Luminometer (Promega)仪器, 按照说明书检测并定量萤火虫荧光素酶和海肾荧光素酶活性。
通过RT-PCR获得油菜ABF2基因的cDNA片段。分析发现, BnaABF2定位于C06染色体, 编码区长度为1 008 bp, 预测蛋白分子量为41.8 kDa。利用BioLinux系统的MEME软件进行motif分析(图1), 发现BnaABF2.1启动子上游包含防御和应激反应、光响应、厌氧诱导、脱落酸响应、水杨酸响应和生长素响应元件。
将BnaABF2蛋白序列与拟南芥、水稻、玉米和油菜ABF蛋白序列进行多重比对, 发现其保守基序具有相似的motif结构(图2), 而转录本结构差异较大, 表明在物种进化过程中ABF的功能具有保守性, 同时不同物种的进化存在差异。因此, 这些基因的功能调控可能在一定程度上存在差异。
为探究油菜BnaABF2的进化情况, 对13个拟南芥ABF、15个水稻ABF、6个油菜ABF、36个小立碗藓(Physcomitrium patens) ABF、22个江南卷柏(Sela-ginella moellendorffii) ABF、7个金牛微球藻(Ostreococcus tauri) ABF以及11个莱茵衣藻(Chlamydomonas reinhardtii) ABF构建了系统进化树。结果显示, 该进化树共有3个较大的分支, BnaABF2与拟南芥ABF2和水稻bZIP23亲缘关系较近(图3)。
转录因子一般在细胞核中行使功能。将BnaABF2的编码区(coding sequence, CDS)构建到融合GFP的pYJHF载体上, 由农杆菌介导瞬时转化本氏烟草。结果显示, 对照GFP信号存在于细胞膜和细胞核中, 而BnaABF2信号集中在细胞核(图4), 表明该转录因子可能在细胞核中发挥作用。
为探究BnaABF2转录因子的基本特性, 通过酵母表达系统检测该转录因子的活性。结果显示, 转化BnaABF2质粒的酵母菌在一缺(SD-W)营养缺陷型培养基上能够生长, 但在二缺(SD-WH+3-AT)以及三缺(SD-WHA)营养缺陷型培养基上无法生长, 并且在X-α-Gal染色实验中变蓝(图5A), 表明BnaABF2在酵母系统中无转录激活活性。
对BnaABF2蛋白序列进行分析, 发现其序列中并不包含EAR抑制性基序(Kagale et al., 2010)。基于这一现象, 推测可能由于密码子的偏好性导致BnaABF2在酵母体内的表达水平偏低。于是, 我们根据低等生物密码子使用上的偏好性对BnaABF2蛋白序列进行密码子优化, 优化后的序列简称为BnaABF2 Co。鉴于发现了BnaABF2蛋白序列中有多个SP/TP位点, 为进一步研究酵母系统中这些位点对BnaABF2转录活性的影响, 我们将预测的BnaABF2的180位、251位、267位、288位和290位丝氨酸(Ser)及270位苏氨酸(Thr)进行定点突变。上述位点突变为丙氨酸(Ala)后的序列简称为BnaABF2 5m (为BnaABF2的非磷酸化失活形式); 突变为天冬氨酸(Asp)后的序列简称为BnaABF2 5D (为BnaABF2的模拟磷酸化突变形式)。将创制好的序列分别构建到pGBKT7载体, 得到重组质粒BD-BnaABF2 Co、BD-BnaABF2 5m以及BD-BnaABF2 5D。我们进一步在酵母系统中检测了这些重组质粒能否激活下游报告基因的正常表达, 结果显示转化BnaABF2 5D质粒的酵母菌在一缺(SD-W)和二缺(SD-WH)营养缺陷型培养基上能够生长, 但是当在二缺营养缺陷型培养基中添加3-AT (SD-WH+3-AT), 或在三缺(SD-WHA)营养缺陷型培养基上, 酵母菌的生长被显著抑制。另外, 在X-α- Gal染色实验中, 这些酵母菌未呈现出蓝色反应,并 且转化了BnaABF2 5m和BnaABF2 Co质粒的酵母菌只能在一缺(SD-W)营养缺陷型培养基上生长(图5B), 表明BnaABF2的密码子优化对其在酵母系统中表现出的转录活性无明显影响, 而我们预测的BnaABF2中的保守SP/TP基序中的Ser/Thr磷酸化位点, 与BnaABF2在酵母系统中的转录活性有关, 并且其模拟磷酸化形式能在一定程度上增强其转录活性。
为探究BnaABF2在不同组织中的表达情况, 对油菜的根、茎、叶、花蕾、花和果荚进行qRT-PCR检测(图6A)。结果显示, BnaABF2在叶中表达量最高, 其次是花, 在茎中最低。为探究BnaABF2在非生物逆境条件下的表达特性, 我们分析了甘蓝型油菜幼苗在不同胁迫(ABA、模拟干旱、热害、盐害和冷害(4°C)处理)下的表达谱, 发现BnaABF2在ABA (50 μmol·L-1)、模拟干旱(15% PEG8000)、热害(38°C)和盐害(200 mmol·L-1 NaCl)胁迫下均有响应。ABA处理后, BnaABF2的表达水平上升, 且处理时间延长后, 转录水平的上升更加显著。模拟干旱胁迫下, BnaABF2的表达水平上升, 转录水平在处理6小时后达到峰值, 随后有所下降, 但仍高于处理1小时后的转录本累积水平。盐胁迫下, BnaABF2的表达水平上升, 但随着盐胁迫时间的延长, 表达水平未表现出显著变化(图6B)。
上述研究表明, BnaABF2转录因子可能在非生物胁迫和植物激素调节方面发挥重要作用。此外, 序列分析表明BnaABF2蛋白中含有多个SP和TP基序, 暗示可能被MPK磷酸化。MPKs作为一种激酶, 能磷酸化下游转录因子, 但MPKs对ABFs的磷酸化却尚未见报道。为探索二者在甘蓝型油菜中的互作关系, 将BnaABF2的编码区克隆到含有黄色荧光蛋白基因的C末端(YFPc), 将BnaMPKs的编码区克隆到含有黄色荧光蛋白基因的N末端(YFPn)双元表达载体上。通过农杆菌介导途径在烟草中瞬时表达。结果显示, 共表达2天后, BnaABF2-YFPc和YFPn-BnaMPK1/2/6/7/9/12/13的组合能够在细胞核中发出黄色荧光信号, 而表达BnaMPK3的组合无任何YFP信号, 说明BnaABF2能与BnaMPK1/2/6/7/9/12/13相互作用(图7)。
MPK激酶一般磷酸化底物蛋白保守的SP/TP位点的S/T氨基酸残基。在BnaABF2蛋白序列中有较多的SP/TP位点, 因此其极有可能被BnaMPKs磷酸化。双分子荧光互补实验表明, 油菜BnaABF2蛋白与BnaMPKs蛋白之间可能存在互作。为进一步解析其互作意义, 我们利用双荧光素酶报告系统检验调控情况。将序列BnaABF2 5m构建入双元表达载体作为效应子, 以BnaABF2的靶基因AtRD29B与萤火虫荧光素酶基因LUC的融合基因作为报告子, 探索BnaABF2的磷酸化失活形式是否影响对下游靶基因的调控。将构建好的重组质粒pBGHF-BnaABF2、pBGHF- BnaABF2 5m、pYJHA-BnaMPK7和pDL-ProAtRD29B共注射到28天龄本氏烟草, 注射后2天进行检测。Danquah等(2015)研究发现, 50 μmol·L-1 ABA处理可促进AtMPK7表达, 因此本研究在收样前4小时向烟草叶片注射50 μmol·L-1 ABA, 以便活化BnaMPK7。结果显示, BnaMPK7对BnaABF2和BnaABF2 5m的作用存在差异, 突变后的BnaABF2对ProAtRD29B启动子下游基因的激活效应减弱, 证明突变的S/T位点中存在BnaMPK7对BnaABF2的磷酸化位点(图8)。
干旱是植物面临的十分常见的一种非生物胁迫。甘蓝型油菜作为重要的油料作物之一, 其理论与应用研究较多。例如, 最近, 李青洋等(2025)对甘蓝型油菜的BnaA02.CPSF6基因进行了克隆及功能分析。我国西北地区的油菜种植受到干旱、盐害以及冷害的影响尤为明显。干旱会导致植物中的ABA水平升高, 进而影响大量胁迫相关基因的表达。这些基因的激活和表达受到转录因子的调控。在ABA介导的基因表达调控中, ABF家族转录因子发挥重要作用(Agarwal and Jha, 2010)。Fujita等(2005)研究表明, 拟南芥ABF2、ABF4和ABF3能被干旱、高盐或ABA处理诱导, 是参与干旱胁迫耐受性的主要转录因子基因, 也参与ABA信号转导。
干旱胁迫下, 拟南芥AREB/ABF三重突变体(areb1/areb2/abf3)的耐受性下降, 同时对外源ABA的敏感性降低。进一步的转录组分析显示, AtABF2、AtABF3和AtABF4基因突变可导致许多渗透胁迫诱导基因的表达水平降低, 表明这些基因在植物响应干旱胁迫中具有关键作用(Yoshida et al., 2010)。它们的功能获得突变体表现出抗旱能力增强(Fujita et al., 2005)。本研究成功克隆了油菜BnaABF2基因的cDNA序列, 通过序列比对发现其蛋白序列在物种间具有保守的motif结构(图2), 但转录本结构存在差异 (图1)。进化树分析显示, 甘蓝型油菜BnaABF2蛋白与拟南芥AtABF2和水稻bZIP23蛋白亲缘关系较近(图3), 其可能具有相似的功能。BnaABF2启动子包含多种胁迫和植物激素响应元件。qRT-PCR结果证明, 该基因在叶中的表达量最高, 且能响应多种逆境和植物激素, 如ABA、模拟干旱以及盐害(图6)。然而, BnaABF2在酵母中未显示出转录激活活性(图5A), 我们初步推测可能是由于密码子的偏好性导致表达水平低。然而, 对BnaABF2蛋白序列的密码子优化后, 仍未显著提升其在酵母中的转录活性(图5B)。进而推测软件预测的磷酸化位点可能与BnaABF2的转录活性密切相关。进一步实验分析, 发现BnaABF2的模拟磷酸化形式的突变型蛋白能有效增强其在酵母系统中的转录活性, 该发现对于正确理解BnaABF2在油菜逆境响应中的调控机制具有一定的借鉴意义。
有研究表明, 干旱和盐胁迫会诱导甘蓝型油菜中BnaABF2基因的表达, 该基因在拟南芥中过表达会增强植株对干旱和盐胁迫的耐受性, 原因是ABF2调控的RD29B、RAB18和KIN2表达及ABA诱导了气孔关闭(Zhao et al., 2016)。水稻中, AREB1同源基因TRAB1能被SnRK2蛋白激酶的同源基因SAPK磷酸化; 小麦中, AREB1同源基因TaABF在体外可以被SnRK2同源基因PKABA1磷酸化(Johnson et al., 2002; Kobayashi et al., 2005)。序列分析与预测发现, 在BnaABF2的氨基酸序列中存在潜在的MPK磷酸化基序, 即保守的SP/TP二肽基序。然而, ABFs是否能被真核生物中广泛存在的MPK磷酸化, 尚未见报道。我们通过荧光双分子互补实验筛选BnaABF2的互作蛋白, 发现BnaABF2能与BnaMPK1/2/6/7/9/12/13互作, 初步验证了这一推测。进一步我们通过双荧光素酶报告系统, 结合点突变的BnaABF2创制, 表明BnaMPK7可能通过磷酸化BnaABF2增强其对下游靶基因的转录调控, 与磷酸化提高了BnaABF2蛋白的稳定性有关。但BnaMPK7与BnaABF2之间的互作、磷酸化及磷酸化的意义还有待进一步验证。
本文对甘蓝型油菜中的逆境响应基因BnaABF2进行研究。启动子分析预测该基因可能响应多种逆境与植物生长调节剂。亚细胞定位结果显示, BnaABF2定位于细胞核。酵母系统转录活性检测表明, Bna-ABF2在酵母系统中无转录激活活性。ABA、模拟干旱以及盐害能够诱导BnaABF2的表达。双分子荧光互补实验证明, BnaABF2蛋白能与BnaMPK1/2/6/7/9/ 12/13蛋白互作, 暗示BnaMPKs可能磷酸化BnaABF2。对BnaABF2潜在的磷酸化位点进行点突变后, 我们通过双荧光素酶报告实验进一步验证了这种可能。总之, 本研究探索了BnaABF2的基本特性及其上游互作MPK蛋白激酶, 可为进一步研究该基因在调控油菜对ABA的应答以及抗旱中的功能与作用机制提供参考借鉴。
江元清: 实验设计与指导, 文章修改; 杨柳卿, 王劲: 开展试验与结果分析, 撰写论文初稿; 燕敬利, 陈芹芹, 程浩坤, 李春, 赵培玉, 杨博: 提供部分实验材料与实验条件。
  • 国家自然科学基金(32171953)
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doi: 10.11983/CBB24019
  • 接收时间:2024-02-05
  • 首发时间:2026-10-27
  • 出版时间:2025-01-10
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  • 收稿日期:2024-02-05
  • 录用日期:2024-08-20
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    西北农林科技大学生命科学学院, 杨凌 712100

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* 江元清, 男, 汉族, 湖南省桃源县人, 教授, 博士生导师。1998年6月本科毕业于湖南农业大学资环学院; 2001年6月硕士研究生毕业于中国农业大学生物学院生物化学与分子生物学专业。2001.07-2003.08在北京市农林科学院农业生物技术研究中心工作(助理研究员), 从事扁桃基因功能分析相关研究。2003.09-2007.11在加拿大阿尔伯特大学(University of Alberta)生物科学系学习(植物生物学专业), 获博士学位。2007.12-2010.11先后在阿尔伯特大学生物科学系和美国加州大学伯克利分校(University of California-Berkeley)植物与微生物学系做博士后研究。2010年12月就职于西北农林科技大学生命科学学院, 从事教学与科研工作。E-mail:
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2种不同金属材料的力学参数
科
Family
属数
Number of
genus
种数
Number of
species
占总种数比例
Percentage of
total species (%)
属
Genus
种数
Number of
species
占总种数比例
Percentage of total
species (%)
鹅膏菌科Amanitaceae 2 11 5.26 鹅膏菌属 Amanita 10 4.78
小菇科 Mycenaceae 2 12 5.74 丝盖伞属 Inocybe 5 2.39
多孔菌科 Polyporaceae 8 14 6.70 蜡蘑属 Laccaria 5 2.39
红菇科 Russulaceae 3 23 11.00 小皮伞属 Marasmius 6 2.87
小菇属 Mycena 11 5.26
光柄菇属 Pluteus 5 2.39
红菇属 Russula 17 8.13
栓菌属 Trametes 5 2.39
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