PwEXP1在青杄种子萌发及逆境响应中的表达特征

doi:10.11707/j.1001-7488.20141208

摘要/Abstract

摘要：

采用RACE-PCR技术,从青杄cDNA文库中克隆得到1个青杄 PwEXP1 基因,其cDNA全长为1 186 bp,含777 bp的完整开放阅读框,编码259个氨基酸.生物信息学分析发现其理论分子量为27.90 kDa,等电点(pI)为8.07,N端有8个半胱氨酸残基,C端4个色氨酸残基,中部1个HFD结构域,具有扩展蛋白的典型结构.RT-qPCR试验显示 PwEXP1 在青杄针叶中相对表达量较高.在种子萌发过程中,未开始萌发的种子表达量较低,萌发第2天和第4天表达量开始升高,萌发6天后达到较高水平.在逆境胁迫下, PwEXP1 在青杄根和叶中的表达模式不同.NaCl和低温、高温胁迫均能增加 PwEXP1 在根中的表达水平; PEG处理的渗透胁迫下, PwEXP1 在根中表达特征呈现前期下调、6 h后上调,而在叶中则在胁迫3 h和6 h时上调、9 h和12 h时下调、24 h时重新上调; 在ABA处理后, PwEXP1 在根和叶中均表现下调.推测 PwEXP1 在青杄种子萌发和逆境响应中发挥作用.

关键词: 青杄, 扩展蛋白, 胁迫响应, 基因表达

Abstract:

The PwEXP1 gene was cloned based on the cDNA library of Picea wilsonii and the EST sequence of PwEXP1 using RACE-PCR method in this study. The full length of PwEXP 1 cDNA was 1 186 bp with an open reading frame (ORF) of 777 bp encoding 259 amino acids. Bioinformatics analysis revealed that the theoretical molecular weight of PwEXP1 was 27.68 kDa with isoelectric point (pI) of 8.07. The protein had a typical structure of the expansin protein: containing 8 conserved Cysteine residues at the N terminal, 4 Tryptophan at the C terminal and a HFD in the middle. With RT-qPCR and RT-PCR the temporal dynamics of expression were conducted. It was found that PwEXP1 gene was expressed in needle, root and stem. In the seed germination process, the expression level of PwEXP1 increased with the germination stage and peaked at 6-10 days. Under different abiotic stress (NaCl, ABA, PEG, low and high temperature), gene expression of PwEXP 1 was investigated by using RT-qPCR in roots and leaves. The expression level of PwEXP 1 was up-regulated in root under NaCl, low and high temperature treatments. Under PEG-induced osmotic stress, the expression of PwEXP1 in roots was down-regulated at the early stage, and then induced after 6 h, while the expression showed an up-down-up trend in leaves. With ABA treatment, the expression of PwEXP1 was slightly repressed in both roots and leaves. The results suggest that PwEXP1 plays important roles in both seed germination and responses to abiotic stresses.

Key words: Picea wilsonii, expansin, stress response, gene expression

中图分类号:

S718.46

张通, 李巧玲, 张凌云. PwEXP1在青杄种子萌发及逆境响应中的表达特征[J]. 林业科学, 2014, 50(12): 56-62.

Zhang Tong, Li Qiaoling, Zhang Lingyun. Expression Characteristics of PwEXP 1 Gene in Seed Germination and Adversity in Picea wilsonii[J]. Scientia Silvae Sinicae, 2014, 50(12): 56-62.

参考文献

荆赞革, 柳李旺, 龚义勤, 等. 2009. 萝卜扩展蛋白基因克隆与表达特征分析. 分子植物育种, 7(4): 801-805.
(Jing Z G, Liu L W, Gong Y Q, et al. 2009. Cloning and expression characterization of the Expansin gene RsEXPB1 in Radish. Molecular Plant Breeding, 7(4): 801-805. [in Chinese] )
李长江, 孙凡, 张通, 等. 2013. 青杄PwPSAF基因的克隆与组织表达分析. 林业科学, 49(10): 40-47.
(Li C J, Sun F, Zhang T, et al. 2013. Cloning and tissue expression analysis of PwPSAF in Picea wilsonii. Scientia Silvae Sinicae, 49(10): 40-47. [in Chinese] )
魏强, 凌雷, 张广忠, 等. 2011. 甘肃兴隆山主要森林类型凋落物累积量及持水特性. 应用生态学报, 22(10): 2589-2598.
(Wei Q, Ling L, Zhang G Z, et al. 2011. Water-holding characteristics and accumulation amount of the litters under main forest types in Xinglong Mountain of Gansu, Northwest China. Chinese Journal of Applied Ecology, 22(10): 2589-2598. [in Chinese] )
徐筱, 黄炳如, 徐吉臣. 2010. 翦股颖AsEXP 1 基因的抗旱性分析. 北京林业大学学报, 32(5): 126-131.
(Xu X, Huang B R, Xu J C. 2010. Analysis of AsEXP1 gene related to drought tolerance in creeping bentgrass. Journal of Beijing Forestry University, 32(5): 126-131. [in Chinese] )
阎伯旭, 曲音波, 高培基. 1998. 色氨酸残基在内切葡聚糖酶分子中的作用. 中国生物化学与分子生物学报, 14(2): 181-185.
(Yan B X, Qu Y B, Gao P J. 1998. Role of tryptophan residue in endoglucanase from Trichoderma pseudokoningii S-38. Chinese Journal of Biochemistry and Molecular Biology, 14(2): 181-185. [in Chinese] )
张盾, 刘亚静, 李长江, 等. 2012. 青杄均一化cDNA文库构建及EST序列分析. 生物技术通报, 8(6): 71-76.
(Zhang D, Liu Y J, Li C J, et al. 2012. Construction of normalized cDNA library and analysis of corresponding EST sequences in Picea wilsonii. Biotechnology Bulletin, 8(6): 71-76. [in Chinese] )
Buchanan C D, Lim S, Salzman R A, et al. 2005. Sorghum bicolor's transcriptome response to dehydration, high salinity and ABA. Plant Mol Biol, 58(5): 699-720.
Chen F, Dahal P, Bradford K J. 2001. Two tomato expansin genes show divergent expression and localization in embryos during seed development and germination. Plant Physiol, 127(3): 928-936.
Choi D, Lee Y, Cho H T, et al. 2003. Regulation of expansin gene expression affects growth and development in transgenic rice plants. Plant Cell, 15(6): 1386-1398.
Dal Santo S, Vannozzi A, Tornielli G B, et al. 2013. Genome-wide analysis of the expansin gene superfamily reveals grapevine-specific structural and functional characteristics. PlosOne, 8(4):e62206. DOI: 10.1371/journal.pone.0062206.
Geilfus C M, Zorb C, Muhling K H. 2010. Salt stress differentially affects growth-mediating beta-expansins in resistant and sensitive maize (Zea mays L.). Plant Physiology and Biochemistry, 48(12): 993-998.
Han Y Y, Li A X, Li F, et al. 2012. Characterization of a wheat (Triticum aestivum L.) expansin gene, TaEXPB 23, involved in the abiotic stress response and phytohormone regulation. Plant Physiology and Biochemistry, 54: 49-58.
Ma J, Li Z, Wang B, et al. 2012. Cloning of an expansin gene from Chimonanthus praecox flowers and its expression in flowers treated with ethephon or 1-methylcyclopropene. Hortscience, 47(10): 1472-1477.
McQueen-Mason S, Durachko D M, Cosgrove D J. 1992. Two endogenous proteins that induce cell wall extension in plants. Plant Cell, 4: 1425-1433.
McQueen-Mason S, Cosgrove D J. 1994. Disruption of hydrogen bonding between plant cell wall polymers by proteins that induce wall extension. Proc Natl Acad Sci USA, 91(14): 6574-6578.
McQueen-Mason S, Cosgrove D J. 1995. Expansin mode of action on cell walls. Analysis of wall hydrolysis, stress relaxation, and binding. Plant Physiol, 107(1): 87-100.
Park C H, Kim T W, Son S H, et al. 2010. Brassinosteroids control AtEXPA 5 gene expression in Arabidopsis thaliana. Phytochemistry, 71(4): 380-387.
Sampedro J, Cosgrove D J. 2005. The expansin superfamily. Genome Biology, 6(12):242.1-242.11.
Sasidharan R, Voesenek L A, Pierik R. 2011. Cell wall modifying proteins mediate plant acclimatization to biotic and abiotic stresses. Critical Reviews in Plant Sciences, 30(6): 548-562.
Sharova E I. 2007. Expansins: Proteins involved in cell wall softening during plant growth and morphogenesis. Russian Journal of Plant Physiology, 54(6): 713-727.
Vreeburg R A, Benschop J J, Peeters A J, et al. 2005. Ethylene regulates fast apoplastic acidification and expansin a transcription during submergence-induced petiole elongation in Rumex palustris. Plant J, 43(4): 597-510.
Wang W, Vinocur B, Altman A. 2003. Plant responses to drought, salinity and extreme temperatures: towards genetic engineering for stress tolerance. Planta, 218(1): 1-14.
Yamada K, Takahashi R, Fujitani C, et al. 2009. Cell wall extensibility and effect of cell-wall-loosening proteins during rose flower opening. Journal of the Japanese Society for Horticultural Science, 78(2): 242-251.
Yu Y, Li Y, Huang G, et al. 2011. PwHAP5, a CCAAT-binding transcription factor, interacts with PwFKBP12 and plays a role in pollen tube growth orientation in Picea wilsonii. J Exp Bot, 62(14): 4805-4817.
Zhao M R, Han Y Y, Feng Y N, et al. 2012. Expansins are involved in cell growth mediated by abscisic acid and indole-3-acetic acid under drought stress in wheat. Plant Cell Reports, 31(4):671-685.

[1]	刘道凤, 王霞, 代银, 杨建峰, 马婧, 李名扬, 眭顺照. 蜡梅转录因子CpTAF10基因的克隆及功能分析[J]. 林业科学, 2019, 55(6): 176-183.
[2]	卢惠君, 李子义, 梁瀚予, 岳远志, 周天畅, 杨玉璋, 王玉成, 及晓宇. 刚毛柽柳NAC24基因的表达及抗逆功能分析[J]. 林业科学, 2019, 55(3): 54-63.
[3]	范伟健, 相伟芳, 王静, 白鹏华, 潘丽娜, 杨艺新, 朱耿平, 李敏. 农药与紫外胁迫对白蛾周氏啮小蜂小分子热激蛋白sHSP12.2基因表达的影响[J]. 林业科学, 2019, 55(2): 128-136.
[4]	张扬, 杜琳, 唐贤丰, 刘唤唤, 周功克, 柴国华. 杨树BAG基因的鉴定及表达模式分析[J]. 林业科学, 2019, 55(1): 138-145.
[5]	魏明科, 俞金健, 黄晓龙, 刘琼瑶, 黄华宏, 林二培, 童再康. 杉木NAC转录因子基因ClNAC1的克隆、表达及单核苷酸多态性分析[J]. 林业科学, 2018, 54(9): 49-59.
[6]	王娟, 陈辉. 华山松大小蠹冷休克结合蛋白的鉴定与表达[J]. 林业科学, 2018, 54(4): 58-66.
[7]	张鹤华, 刘嘉欣, 罗朝兵, 张凌云. 青杄转录因子PwERF8及其启动子序列的克隆与分析[J]. 林业科学, 2018, 54(3): 48-60.
[8]	李建波, 贾会霞, 张进, 刘伯斌, 胡建军, 王丽娟, 卢孟柱. 毛白杨PtoWOX4a基因过表达对次生生长的影响[J]. 林业科学, 2018, 54(2): 52-59.
[9]	刘长英, 张梦, 魏从进, 许亚珍, 曹博宁, 郑莎, 范伟, 向仲怀, 赵爱春. 转桑树MAPK5基因拟南芥在不同逆境胁迫下的抗性分析[J]. 林业科学, 2017, 53(9): 35-44.
[10]	孙化雨, 李利超, 赵韩生, 杨意宏, 王思宁, 高志民. 黄藤2个NAC基因的分子特征及其SSR分子标记开发[J]. 林业科学, 2017, 53(8): 132-140.
[11]	唐绯绯, 赵玉琳, 王培龙, 冯德明, 宋怡, 高彩球. 刚毛柽柳ThP5CR基因的克隆及抗逆功能分析[J]. 林业科学, 2017, 53(7): 1-9.
[12]	李楠, 郑勇奇, 丁红梅, 柳新红, 盛炜彤, 江波, 李海波. 低温胁迫下短枝木麻黄耐寒相关基因的差异表达分析[J]. 林业科学, 2017, 53(7): 62-71.
[13]	游韩莉, 袁义杭, 李长江, 张凌云. 青杄MYB转录因子基因PwMYB20的克隆及表达分析[J]. 林业科学, 2017, 53(5): 23-32.
[14]	余建, 赵爱春, 刘长英, 梁燕梅, 朱攀攀, 蔡雨翔, 王茜龄, 李镇刚, 余茂德. 外源乙烯利与1-MCP处理对桑椹中乙烯和花青素相关代谢基因表达的影响[J]. 林业科学, 2017, 53(2): 138-148.
[15]	王凯利, 傅鹰, 周明兵. 毛竹SQUAMOSA启动子结合蛋白SBP家族基因的鉴定及表达模式分析[J]. 林业科学, 2017, 53(12): 50-61.