84K杨4CL3/4CL5基因克隆及生物信息学分析

doi:10.7525/j.issn.1673-5102.2019.04.009

植物研究 ›› 2019, Vol. 39 ›› Issue (4): 547-556.doi: 10.7525/j.issn.1673-5102.2019.04.009

84K杨4CL3/4CL5基因克隆及生物信息学分析

孙晓莎, 王遂, 赵曦阳, 曲冠证

林木遗传育种国家重点实验室, 东北林业大学, 哈尔滨 150040

收稿日期:2019-01-02 出版日期:2019-07-05 发布日期:2019-07-03
通讯作者: 曲冠证 E-mail:quguanzheng@yahoo.com
作者简介:孙晓莎(1991-),女,硕士研究生,主要从事林木遗传育种研究。
基金资助:
国家自然科学基金面上项目（31500549，31670675）

Cloning and Bioinformatics Analysis 4CL3/4CL5 Gene of Populus alba×P.glandulosa

SUN Xiao-Sha, WANG Sui, ZHAO Xi-Yang, QU Guan-Zheng

State Key Laboratory of Tree Genetics and Breeding, Northeast Forestry University, Harbin 150040

Received:2019-01-02 Online:2019-07-05 Published:2019-07-03
Supported by:
National Natural Science Foundation of China(31500549,31670675)

摘要/Abstract

摘要： 木质素是木材的重要组成成分，主要起机械支持、水分运输及防御病虫害等作用。4CL酶是控制木质素合成途径中的关键酶之一。利用PCR技术从84K杨幼叶cDNA中克隆得到Pag4CL3/4CL5基因，GenBank 登录号分别为MK183033（Pag4CL3）及MK183034（Pag4CL5）。利用生物信息学软件，对这两个基因的功能和特征进行分析。结果发现84K杨中4CL3/4CL5蛋白与毛果杨中4CL3/4CL5蛋白相似度高达97%；此外，均含有SSGTTGLPKGV和GEICIRG两个保守基序；亚细胞定位预测显示Pag4CL3/4CL5蛋白主要定位在内质网中，推测可能是一种膜蛋白；蛋白的亲水性预测Pag4CL3/4CL5均为亲水性蛋白。通过qRT-PCR分析Pag4CL3/4CL5在不同组织部位中的表达差异，结果发现Pag4CL3在叶及茎中表达量较高，在根和顶芽中表达量较低；Pag4CL5在叶及根中表达量较高，在茎和顶芽中表达量较低，两个同源基因表达具有组织部位的差异性。Pag4CL3和Pag4CL5可能在叶中共同行使功能，Pag4CL5主要在根中行使功能，Pag4CL3主要在茎中行使功能。

关键词: 84K杨, 4CL基因, 克隆, 生物信息学分析

Abstract: Lignin is an important component of wood, mainly for mechanical support, water transport and defense against pests and diseases. The 4CL enzyme is one of the key enzymes in controlling the pathway of lignin synthesis. The Pag4CL3/4CL5 gene was cloned from 84K poplar leaf cDNA by PCR. GenBank accession numbers are MK183033(Pag4CL3) and MK183034(Pag4CL5). The functions and characteristics of the two genes were analyzed using bioinformatics software. The similarity of the 4CL3/4CL5 proteins in 84K poplar and the 4CL3/4CL5 proteins in Populus trichocarpa were 97%,and they contained two conserved motifs of SSGTTGLPKGV and GEICIRG; subcellular localization showed the main localization of Pag4CL3/4CL5 proteins. In the endoplasmic reticulum, it may be a membrane protein and both are hydrophilic proteins. The expression of the Pag4CL3/4CL5 gene in different tissues was analyzed by qRT-PCR. The results showed that the Pag4CL3 gene was mainly expressed in leave and stem, and the expression level in root and top buds was low. The Pag4CL5 gene showed high expression level in leave and root, and low level in stem and apical buds. Pag4CL3 and Pag4CL5 may function together in the leaves, however, Pag4CL5 ismainly function in the root, and Pag4CL3 mainly function in the stems.

Key words: Populus alba×P. glandulosa, 4CL gene, cloning, bioinformatics analysis

中图分类号:

S792.119

孙晓莎, 王遂, 赵曦阳, 曲冠证. 84K杨4CL3/4CL5基因克隆及生物信息学分析[J]. 植物研究, 2019, 39(4): 547-556.

SUN Xiao-Sha, WANG Sui, ZHAO Xi-Yang, QU Guan-Zheng. Cloning and Bioinformatics Analysis 4CL3/4CL5 Gene of Populus alba×P.glandulosa[J]. Bulletin of Botanical Research, 2019, 39(4): 547-556.

参考文献

1. Hu W J,Harding S A,Lung J,et al. Repression of lignin biosynthesis promotes cellulose accumulation and growth in transgenic trees[J]. Nature Biotechnology,1999,17(8):808-812.
2. Sarkanen K V,Ludwig C H. Lignins,occurrence,formation,structure and reactions[M]. New York:Wiley-Interscience,1971:95-163.
3. Chiang V L,Puumala R J,Takeuchi H,et al. Comparison of softwood and hardwood kraft pulping[J]. Tappi Journal,1988,71(9):173-176.
4. Hawkins S W,Boudet A M. Purification and characterization of cinnamyl alcohol dehydrogenase isoforms from the periderm of Eucalyptus gunnii Hook[J]. Plant Physiology,1994,104(1):75-84.
5. Matsui N,Chen F,Yasuda S,et al. Conversion of guaiacyl to syringyl moieties on the cinnamyl alcohol pathway during the biosynthesis of lignin in angiosperms[J]. Planta,2000,210(5):831-835.
6. Li Q Z,Song J,Peng S B,et al. Plant biotechnology for lignocellulosic biofuel production[J]. Plant Biotechnology Journal,2014,12(9):1174-1192.
7. 吴锦程,唐朝晖,陈群,等. 不同贮藏温度对枇杷果肉木质化及相关酶活性的影响[J]. 植物科学学报,2006,24(3):235-239.
Wu J C,Tang C H,Chen Q,et al. The effects of different storage temperatures on lignification and related enzymes activities in loquat pulp[J]. Plant Science Journal,2006,24(3):235-239.
8. 李慧,赵慧,闵现东,等. 84K杨引种栽培及扦插育苗研究[J]. 现代农业科技,2017,(13):146-147.
Li H,Zhao H,Yan X D,et al. Introduction and cultivation of 84K poplar and cutting seedlings[J]. Modern Journal of Agricultural Science and Technology,2017(13):146-147.
9. Challis G L,Ravel J,Townsend C A. Predictive,structure-based model of amino acid recognition by nonribosomal peptide synthetase adenylation domains[J]. Chemistry & Biology,2000,7(3):211-224.
10. Fulda M,Heinz E,Wolter F P. The fadD gene of Escherichia coli K12 is located close to rnd at 39.6 min of the chromosomal -map and is a new member of the AMP-binding protein family[J]. Molecular and General Genetics MGG,1994,242(3):241-249.
11. Whetten R W,Mackay J J,Sederoff R R. Recent advances in understanding lignin biosynthesis[J]. Annual Review of Plant Physiology and Plant Molecular Biology,1998,49(1):585-609.
12. Costa M A,Bedgar D L,Moinuddin S G A,et al. Characterization in vitro and in vivo of the putative multigene 4-coumarate:CoA ligase network in Arabidopsis:syringyl lignin and sinapate/sinapyl alcohol derivative formation[J]. Phytochemistry,2005,66(17):2072-2091.
13. Soltani B M,Ehlting J,Hamberger B,et al. Multiple cis-regulatory elements regulate distinct and complex patterns of developmental and wound-induced expression of Arabidopsis thaliana 4CL gene family members[J]. Planta,2006,224(5):1226-1238.
14. Voo K S,Whetten R W,O'malley D M,et al. 4-coumarate:coenzyme a ligase from loblolly pine xylem. Isolation,characterization,and complementary DNA cloning[J]. Plant Physiology,1995,108(1):85-97.
15. Chowdhury M E K,Choi B,Cho B K,et al. Regulation of 4CL,encoding 4-coumarate:coenzyme a ligase,expression in kenaf under diverse stress conditions[J]. Plant Omics Journal,2013,6(4):254-262.
16. Hu W J,Kawaoka A,Tsai C J,et al. Compartmentalized expression of two structurally and functionally distinct 4-coumarate:CoA ligase genes in aspen(Populus tremuloides)[J]. Proceedings of the National Academy of Sciences of the United States of America,1998,95(9):5407-5412.
17. 饶国栋,张永卓,魏弘宜,等. 毛白杨4-香豆酸:辅酶A连接酶(4CL3)的酶学特征研究[J]. 林业科学究,2013,26(5):542-547.
Rao G D,Zhang Y Z,Wei H Y,et al. Enzymatic characteristics of 4-coumarate:coenzyme a ligase(4CL3) from Populus tomentosa[J]. Forest Research,2013,26(5):542-547.
18. Lois R,Hahlbrock K. Differential wound activation of members of the phenylalanine ammonia-lyase and 4-coumarate:CoA ligase gene families in various organs of parsley plants[J]. Zeitschrift für Naturforschung C,1992,47(1-2):90-94.
19. Ehlting J,Büttner D,Wang Q,et al. Three 4-coumarate:coenzyme A ligases in Arabidopsis thaliana represent two evolutionarily divergent classes in angiosperms[J]. Plant Journal,1999,19(1):9-20.

[1]	张昊楠, 陈珊珊, 徐建民, 罗萍, 王晓萍, 许志茹, 范春节. 巨桉EgrWAT1基因克隆和功能初步分析[J]. 植物研究, 2023, 43(4): 601-611.
[2]	宋海云, 张涛, 贺鹏, 郑树芳, 王立丰, 王文林. 澳洲坚果MibZIP1基因克隆及表达规律分析[J]. 植物研究, 2023, 43(1): 131-139.
[3]	李娜, 王潇楠. 拟南芥种皮色素形成突变体的筛选与表型鉴定[J]. 植物研究, 2023, 43(1): 59-68.
[4]	李登高, 林睿, 穆青慧, 周娜, 张焱如, 白薇. 马铃薯StCRKs基因家族的鉴定分析及响应逆境信号的表达[J]. 植物研究, 2022, 42(6): 1033-1043.
[5]	李登高, 林睿, 穆青慧, 周娜, 张焱如, 白薇. 马铃薯StNPR4基因的克隆与功能分析[J]. 植物研究, 2022, 42(5): 821-829.
[6]	栗赫铭, 秦洪涛, 魏德强, 李旭, 蓝兴国. 羽衣甘蓝BoMAPK4原核表达、抗体制备及蛋白表达分析[J]. 植物研究, 2022, 42(4): 584-591.
[7]	王雪莹, 王瑞琪, 张洋, 刘聪, 夏德安, 魏志刚. 毛果杨CNGC家族全基因组鉴定及胁迫响应分析[J]. 植物研究, 2022, 42(4): 613-625.
[8]	程赫, 田双慧, 张洋, 刘聪, 夏德安, 魏志刚. 毛果杨nsLTP基因家族全基因组水平鉴定及其表达特性分析[J]. 植物研究, 2022, 42(3): 412-423.
[9]	杨宇宁, 董昊, 董实伟, 王乃锐, 宋跃, 张含国, 李淑娟. 长白落叶松转录因子LobHLH34克隆及表达分析[J]. 植物研究, 2022, 42(1): 112-120.
[10]	赵佳明, 樊二勤, 刘轶, 王智, 王军辉, 曲冠证. 楸树CbuATX1，CbuATX1-like和CbuATX2基因克隆及生物信息学分析[J]. 植物研究, 2022, 42(1): 47-61.
[11]	谭秋锦, 张涛, 韦媛荣, 郑树芳, 汤秀华, 王文林. 澳洲坚果开花相关MiMADS-box基因克隆及表达载体构建[J]. 植物研究, 2021, 41(6): 888-895.
[12]	纪欣童, 于磊, 詹亚光. 水曲柳BZR1基因克隆及其表达模式分析[J]. 植物研究, 2021, 41(5): 744-752.
[13]	柯维忠, 钟雯娟, 刘凯盈, 李祥媛, 尹明华. 黄独微型块茎鲨烯合酶的基因克隆与序列分析[J]. 植物研究, 2021, 41(2): 243-250.
[14]	彭淑萍, 董诚明, 朱畇昊. 响应内生菌侵染的两个地黄茉莉酸合成关键基因的克隆与表达分析[J]. 植物研究, 2021, 41(2): 294-301.
[15]	王文林, 陈海生, 郑树芳, 樊松乐, 王立丰, 谭秋锦, 覃振师, 黄锡云, 贺鹏, 汤秀华, 许鹏. 澳洲坚果MiMYB2基因克隆及结构与功能分析[J]. 植物研究, 2020, 40(6): 913-922.

84K杨4CL3/4CL5基因克隆及生物信息学分析

Cloning and Bioinformatics Analysis 4CL3/4CL5 Gene of Populus alba×P.glandulosa

PDF (PC)

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

编辑推荐

Metrics

本文评价