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毛竹PhebHLH6基因克隆及表达分析

卓娟 侯丹 林新春

卓娟, 侯丹, 林新春. 毛竹PhebHLH6基因克隆及表达分析[J]. 浙江农林大学学报. doi: 10.11833/j.issn.2095-0756.20220553
引用本文: 卓娟, 侯丹, 林新春. 毛竹PhebHLH6基因克隆及表达分析[J]. 浙江农林大学学报. doi: 10.11833/j.issn.2095-0756.20220553
ZHUO Juan, HOU Dan, LIN Xinchun. Cloning and expression analysis of bHLH6 gene from Phyllostachys edulis[J]. Journal of Zhejiang A&F University. doi: 10.11833/j.issn.2095-0756.20220553
Citation: ZHUO Juan, HOU Dan, LIN Xinchun. Cloning and expression analysis of bHLH6 gene from Phyllostachys edulis[J]. Journal of Zhejiang A&F University. doi: 10.11833/j.issn.2095-0756.20220553

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毛竹PhebHLH6基因克隆及表达分析

doi: 10.11833/j.issn.2095-0756.20220553
基金项目: 浙江省自然科学基金重点资助项目(LZ20C160002);国家自然科学基金资助项目(32150410354,31971735)
详细信息
    作者简介: 卓娟(ORCID: 0000-0003-4283-7848),从事植物生物技术等研究。E-mail: zj9597v@163.com
    通信作者: 林新春(ORCID: 0000-0001-9508-526X),教授,博士,从事植物生物技术等研究。E-mail: linxcx@163.com
  • 中图分类号: S722.3

Cloning and expression analysis of bHLH6 gene from Phyllostachys edulis

  • 摘要:   目的  研究PhebHLH6转录因子在毛竹Phyllostachys edulis逆境胁迫应答中的作用,为毛竹抗逆分子机制研究奠定一定的基础。  方法  以毛竹实生苗为材料进行非生物胁迫处理[干旱胁迫、盐胁迫、水杨酸(SA)和脱落酸(ABA)处理],利用转录组数据筛选出1条差异表达基因,命名为PhebHLH6,并对其进行了基因克隆及生物信息学分析;采用实时荧光定量PCR方法分析PhebHLH6在干旱、盐胁迫及SA、ABA处理下的表达模式。  结果  PhebHLH6基因编码区长度为801 bp,编码266个氨基酸,包含bHLH结构域,属于典型的bHLH转录因子。组织特异性表达分析表明:PhebHLH6在毛竹各个组织均有表达,其中在1.5和3.0 m的笋顶部表达丰度最高。在干旱和高盐胁迫处理下,PhebHLH6的表达水平在处理3 h时被强烈诱导,但在处理24 h后显著下调。在SA和ABA激素处理下,PhebHLH6的表达水平被SA和ABA 诱导也呈先上升再下降的趋势,其中受SA强烈诱导,受ABA诱导作用较弱。  结论  PhebHLH6可能参与了毛竹干旱和盐胁迫早期响应途径,并可能在SA和ABA激素信号通路中起一定的调控作用。图4表2参32
  • 图  1  不同物种bHLH6基因氨基酸序列比对

    Figure  1  Alignment of bHLH6 amino acid sequences from different species

    图  2  不同物种bHLH6-like氨基酸序列进化树构建

    Figure  2  Phylogenetic tree construction of BHLH6-like amino acid sequences

    图  3  毛竹不同组织中PhebHLH6基因的表达

    Figure  3  Expression level of PhebHLH6 gene in different tissues

    图  4  非生物胁迫以及激素处理下PhebHLH6表达模式

    Figure  4  Expression patterns of PhebHLH6 under abiotic stress and hormone treatment

    表  1  基因克隆及表达所用引物序列

    Table  1.   Primers used in gene clone and quantitative real-time PCR

    用途引物名称序列(5′→3′)
    基因克隆 PhebHLH6-F ATGGACGCGGACATGGGCGACA
    PhebHLH6-R CTAATAGCTCATCGAGCTCGGG
    GGGCTTC
    实时荧光定量
    PCR (RT-qPCR)
    Q-PhebHLH6-F CGAGAAGCTATACGCGATCC
    Q-PhebHLH6-F CTGCAGCTGCTGGATGTAAT
    Q-NTB-F TCTTGTTTGACACCGAAGAGGAG
    Q-NTB-F AATAGCTGTCCCTGGAGGAGTTT
    下载: 导出CSV

    表  2  PhebHLH6基因启动子顺式作用元件分析

    Table  2.   Cis-element analysis of PhebHLH6 gene promoter

    作用元件序列数量功能作用元件序列数量功能
    ABRE CACGTG 9 脱落酸响应元件 Sp1 GGGCGG 1 光响应元件
    ARE AAACCA 1 厌氧诱导顺式作用元件 chs-CMA2a TCACTTGA 1 光响应元件
    CAAT-box CCAAT 15 启动子和增强子区域调控元件 TGA-element AACGAC 2 生长素响应元件
    CGTCA-motif CGTCA 5 茉莉酸甲酯响应元件 O2-site GATGATGTGG 1 玉米醇溶蛋白代谢调节元件
    MRE AACCTAA 1 光响应元件 TATA-box TATA 35 核心启动子元件
    G-Box TACGTG 3 光响应元件 TGACG-motif TGACG 5 茉莉酸甲酯响应元件
    下载: 导出CSV
  • [1] PENG Zhenhua, LU Ying, LI Lubin, et al. The draft genome of the fast-growing non-timber forest species moso bamboo (Phyllostachys heterocycla) [J]. Nature Genetics, 2013, 45(4): 456 − 461. doi:  10.1038/ng.2569
    [2] 胡智勇. 毛竹的生物学特性及栽植技术[J]. 安徽农学通报, 2014, 20(12): 117 − 118. doi:  10.16377/j.cnki.issn1007-7731.2014.12.058

    HU Zhiyong. Biological characteristics and planting techniques of Phyllostachys edulis [J]. Anhui Agricultural Science Bulletin, 2014, 20(12): 117 − 118. doi:  10.16377/j.cnki.issn1007-7731.2014.12.058
    [3] 徐秀荣, 杨克彬, 王思宁, 等. 毛竹bHLH转录因子的鉴定及其在干旱和盐胁迫条件下的表达分析[J]. 植物科学学报, 2019, 37(5): 610 − 620. doi:  10.11913/PSJ.2095-0837.2019.50610

    XU Xiurong, YANG Kebin, WANG Sining, et al. Identification of bHLH transcription factors in moso bamboo (Phyllostachys edulis) and their expression analysis under drought and salt stress [J]. Plant Science Journal, 2019, 37(5): 610 − 620. doi:  10.11913/PSJ.2095-0837.2019.50610
    [4] 吕玉龙. 高温干旱对毛竹林的危害及抗旱经营措施建议[J]. 林业实用技术, 2014(8): 53 − 55.

    LÜ Yulong. Harm of high temperature and drought on moso bamboo forest and suggestions on drought resistance management [J]. Practical Forestry Technology, 2014(8): 53 − 55.
    [5] 毛美红, 丁笑章, 傅柳方, 等. 干旱对毛竹林新竹成竹影响的调查分析[J]. 世界竹藤通讯, 2012, 10(1): 12 − 15. doi:  10.3969/j.issn.1672-0431.2012.01.007

    MAO Meihong, DING Xiaozhang, FU Liufang, et al. Investigation of the effect of drought on new moso forest cultivation [J]. World Bamboo and Rattan, 2012, 10(1): 12 − 15. doi:  10.3969/j.issn.1672-0431.2012.01.007
    [6] CHEN Yiyun, LI Mengyao, WU Xuejun, et al. Genome-wide analysis of basic helix-loop-helix family transcription factors and their role in responses to abiotic stress in carrot [J]. Molecular Breeding, 2015, 35(5): 1 − 12. doi:  10.1007/s11032-015-0319-0
    [7] SONNENFELD M J, DELVECCHIO C, SUN Xuetao. Analysis of the transcriptional activation domain of the Drosophila tango bHLH-PAS transcription factor [J]. Development Genes and Evolution, 2005, 215(5): 221 − 229. doi:  10.1007/s00427-004-0462-9
    [8] BAILEY P C, MARTIN C, TOLEDO-ORTIZ G, et al. Update on the basic helix-loop-helix transcription factor gene family in Arabidopsis thaliana [J]. The Plant Cell, 2003, 15(11): 2497 − 2502. doi:  10.1105/tpc.151140
    [9] LI Xiaoqing, TANG Yuanping, YUAN Zheng, et al. Genome-wide analysis of basic/helix-loop-helix transcription factor family in rice and Arabidopsis [J]. Plant Physiology, 2006, 141(4): 1167 − 1184. doi:  10.1104/pp.106.080580
    [10] LEDENT V, VERVOORT M. The basic helix-loop-helix protein family: comparative genomics and phylogenetic analysis [J]. Genome Research, 2001, 11(5): 754 − 770. doi:  10.1101/gr.177001
    [11] RIECHMANN J L, HEARD J E, MARTIN G, et al. Arabidopsis transcription factors: genome-wide comparative analysis among eukaryotes [J]. Science, 2000, 290(5499): 2105 − 2110. doi:  10.1126/science.290.5499.2105
    [12] AN Jianping, LI Haohao, SONG Laiqing, et al. The molecular cloning and functional characterization of MdMYC2, a bHLH transcription factor in apple [J]. Plant Physiology and Biochemistry, 2016, 108: 24 − 31. doi:  10.1016/j.plaphy.2016.06.032
    [13] KONDOU Y, NAKAZAWA M, KAWASHIMA M, et al. RETARDED GROWTH OF EMBRYO1, a new basic helix-loop-helix protein, expresses in endosperm to control embryo growth [J]. Plant Physiology, 2008, 147(4): 1924 − 1935. doi:  10.1104/pp.108.118364
    [14] HEISLER M G, ATKINSON A, BYLSTRA Y H, et al. SPATULA, a gene that controls development of carpel margin tissues in Arabidopsis, encodes a bHLH protein [J]. Development, 2001, 128(7): 1089 − 1098. doi:  10.1242/dev.128.7.1089
    [15] WANG Houping, LI Yang, PAN Jinjing, et al. The bHLH transcription factors MYC2, MYC3, and MYC4 are required for jasmonate-mediated inhibition of flowering in Arabidopsis [J]. Molecular Plant, 2017, 10(11): 1461 − 1464.
    [16] TOLEDO-ORTIZ G, HUQ E, QUAIL P H. The Arabidopsis basic/helix-loop-helix transcription factor family [J]. Plant Cell, 2003, 15(8): 1749 − 1770. doi:  10.1105/tpc.013839
    [17] HEIM M A, JAKOBY M, WERBER M, et al. The basic helix-loop-helix transcription factor family in plants: a genome-wide study of protein structure and functional diversity[J]. Molecular Biology and Evolution, 20(5): 735 − 747.
    [18] FAN Yu, YANG Hao, LAI Dili, et al. Genome-wide identification and expression analysis of the bHLH transcription factor family and its response to abiotic stress in sorghum [Sorghum bicolor (L. ) Moench] [J/OL]. BMC Genomics, 2021, 22: 415[2022-07-30]. doi:  10.1186/s12864-021-07652-9.
    [19] ZHAO Hansheng, GAO Zhimin, WANG Le, et al. Chromosome-level reference genome and alternative splicing atlas of moso bamboo (Phyllostachys edulis) [J/OL]. GigaScience, 2018, 7(10): giy115[2022-07-30]. doi:  10.1093/gigascience/giy115.
    [20] RYCHLIK W. OLIGO 7 primer analysis software [J]. PCR Primer Design, 2007, 402: 35 − 59.
    [21] GUO Zhenhua, MA Pengfei, YANG Guoqian, et al. Genome sequences provide insights into the reticulate origin and unique traits of woody bamboos [J]. Molecular Plant, 2019, 12(10): 1353 − 1365.
    [22] CHEN Chengjie, CHEN Hao, ZHANG Yi, et al. TBtools: an integrative toolkit developed for interactive analyses of big biological data [J]. Molecular Plant, 2020, 13(8): 1194 − 1202.
    [23] LIVAK K J, SCHMITTGEN T D. Analysis of relative gene expression data using real-time quantitative PCR and the $2^{-\Delta\Delta C_{T}} $ method[J]. Methods, 2000, 25(4): 402-408.
    [24] FAN Chunjie, MA Jinmin, GUO Qirong, et al. Selection of reference genes for quantitative real-time PCR in bamboo (Phyllostachys edulis) [J/OL]. PLoS ONE, 2013, 8(2): e56573[2022-07-31]. doi: 10.1371/journal. pone. 0056573.
    [25] MAO Ke, DONG Qinglong, LI Chao, et al. Genome wide identification and characterization of apple bHLH transcription factors and expression analysis in response to drought and salt stress[J/OL]. Front in Plant Science, 2017, 8: 480[2022-07-30]. doi:  10.3389/fpls.2017.00480.
    [26] HE Qiuju, LU Hong, GUO Huaxing, et al. OsbHLH6 interacts with OsSPX4 and regulates the phosphate starvation response in rice [J]. The Plant Journal, 2020, 105(3): 649 − 667.
    [27] 张子佳, 王迪, 傅彬英. 水稻转录因子bHLH家族基因响应环境胁迫表达谱分析[J]. 分子植物育种, 2008, 6(3): 425 − 431. doi:  10.3969/j.issn.1672-416X.2008.03.003

    ZHANG Zijia, WANG Di, FU Binying. Expression patterns of rice bHLH genes responsive to environmental stresses [J]. Molecular Plant Breeding, 2008, 6(3): 425 − 431. doi:  10.3969/j.issn.1672-416X.2008.03.003
    [28] 李朝霞, 高强, 刘雅正, 等. 玉米 ZmPTF1 基因克隆和过表达分析[J]. 湖南农业大学学报(自然科学版), 2007, 33(1): 92 − 96.

    LI Zhaoxia, GAO Qiang, LIU Yazheng, et al. Cloning of ZmPTF1 from Zea mays and its over expression analysis [J]. Journal of Hunan Agricultural University (Natural Sciences), 2007, 33(1): 92 − 96.
    [29] MENG Fanwei, YANG Chao, CAO Jidong, et al. A bHLH transcription activator regulates defense signaling by nucleo-cytosolic trafficking in rice [J]. Journal of Integrative Plant Biology, 2020, 62(10): 1552 − 1573. doi:  10.1111/jipb.12922
    [30] WU Hua, YE Haiyan, YAO Ruifeng, et al. OsJAZ9 acts as a transcriptional regulator in jasmonate signaling and modulates salt stress tolerance in rice [J]. Plant Science, 2015, 232: 1 − 12. doi:  10.1016/j.plantsci.2014.12.010
    [31] KIRIBUCHI K, SUGIMORI M, TAKEDA M, et al. RERJ1, a jasmonic acid-responsive gene from rice, encodes a basic helix-loop-helix protein [J]. Biochemical and Biophysical Research Communications, 2004, 325(3): 857 − 863. doi:  10.1016/j.bbrc.2004.10.126
    [32] KIRIBUCHI K, JIKUMARU Y, KAKU H, et al. Involvement of the basic helix-loop-helix transcription factor RERJ1 in wounding and drought stress responses in rice plants [J]. Bioscience,Biotechnology,and Biochemistry, 2005, 69(5): 1042 − 1044. doi:  10.1271/bbb.69.1042
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  • 收稿日期:  2022-08-30
  • 修回日期:  2022-12-14
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毛竹PhebHLH6基因克隆及表达分析

doi: 10.11833/j.issn.2095-0756.20220553
    基金项目:  浙江省自然科学基金重点资助项目(LZ20C160002);国家自然科学基金资助项目(32150410354,31971735)
    作者简介:

    卓娟(ORCID: 0000-0003-4283-7848),从事植物生物技术等研究。E-mail: zj9597v@163.com

    通信作者: 林新春(ORCID: 0000-0001-9508-526X),教授,博士,从事植物生物技术等研究。E-mail: linxcx@163.com
  • 中图分类号: S722.3

摘要:   目的  研究PhebHLH6转录因子在毛竹Phyllostachys edulis逆境胁迫应答中的作用,为毛竹抗逆分子机制研究奠定一定的基础。  方法  以毛竹实生苗为材料进行非生物胁迫处理[干旱胁迫、盐胁迫、水杨酸(SA)和脱落酸(ABA)处理],利用转录组数据筛选出1条差异表达基因,命名为PhebHLH6,并对其进行了基因克隆及生物信息学分析;采用实时荧光定量PCR方法分析PhebHLH6在干旱、盐胁迫及SA、ABA处理下的表达模式。  结果  PhebHLH6基因编码区长度为801 bp,编码266个氨基酸,包含bHLH结构域,属于典型的bHLH转录因子。组织特异性表达分析表明:PhebHLH6在毛竹各个组织均有表达,其中在1.5和3.0 m的笋顶部表达丰度最高。在干旱和高盐胁迫处理下,PhebHLH6的表达水平在处理3 h时被强烈诱导,但在处理24 h后显著下调。在SA和ABA激素处理下,PhebHLH6的表达水平被SA和ABA 诱导也呈先上升再下降的趋势,其中受SA强烈诱导,受ABA诱导作用较弱。  结论  PhebHLH6可能参与了毛竹干旱和盐胁迫早期响应途径,并可能在SA和ABA激素信号通路中起一定的调控作用。图4表2参32

English Abstract

卓娟, 侯丹, 林新春. 毛竹PhebHLH6基因克隆及表达分析[J]. 浙江农林大学学报. doi: 10.11833/j.issn.2095-0756.20220553
引用本文: 卓娟, 侯丹, 林新春. 毛竹PhebHLH6基因克隆及表达分析[J]. 浙江农林大学学报. doi: 10.11833/j.issn.2095-0756.20220553
ZHUO Juan, HOU Dan, LIN Xinchun. Cloning and expression analysis of bHLH6 gene from Phyllostachys edulis[J]. Journal of Zhejiang A&F University. doi: 10.11833/j.issn.2095-0756.20220553
Citation: ZHUO Juan, HOU Dan, LIN Xinchun. Cloning and expression analysis of bHLH6 gene from Phyllostachys edulis[J]. Journal of Zhejiang A&F University. doi: 10.11833/j.issn.2095-0756.20220553

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