三个烟草防御素基因的克隆及序列分析

林世锋; 任学良; 王轶; 王仁刚

doi:10.3969/j.issn.1007-5119.2012.05.004

三个烟草防御素基因的克隆及序列分析

贵州省烟草科学研究所, 贵阳 550081

基金项目: 贵州省科技厅农业攻关项目(2011-3047);贵州省烟草专卖局资助项目(黔烟科2009-11);贵州省科学技术基金资助项目{黔科合 J字[2012]2256号}

详细信息

作者简介:
林世锋, 男, 博士, 主要从事烟草遗传育种与分子生物学研究。E-mail:linshifeng1978@163.com

通讯作者:
王仁刚

中图分类号: S572.01
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出版历程
- 收稿日期: 2011-12-15
- 修回日期: 2012-03-18
- 刊出日期: 2012-10-29

In Silicon Cloning and Sequence Analysis of Three Defensin Genes in Tobacco (Nicotiana tabacum)

Guizhou Tobacco Research Institute, Guiyang 550081, China

摘要

摘要: 为从烟草中克隆与植物抗病虫反应相关的防御素基因, 以辣椒防御素基因为探针, 通过电子克隆的方法从烟草栽培品种K326中克隆到3个防御素基因(NtDEF1、NtDEF2和NtDEF3), 并对其进行了序列分析。结果表明, 3个防御素cDNA序列均包含完整的开放读码框, 分别编码78、77、76个氨基酸, 具有8个半胱氨酸的高度保守结构等典型的植物防御素特征, 成熟肽含有γ-硫素蛋白功能结构域。通过同源比对和进化分析发现, NtDEF1、NtDEF2和NtDEF3的氨基酸序列与辣椒的一致性最高为50%~87%, 亲缘关系较近;但与已报道的烟草属植物防御素的一致性仅为33%~40%, 亲缘关系较远, 表明NtDEF1、NtDEF2和NtDEF3是烟草中未报道的3个防御素基因。
- 烟草 /
- 防御素 /
- 克隆 /
- 序列分析
Abstract: In silicon cloning, three distinct defensin genes (NtDEF1, NtDEF2 and NtDEF3) were obtained successfully from tobacco cultivar K326 by using pepper CaDEF1 gene sequence (AF442388) to be probe. Sequence analysis showed that the in silicon cloned cDNAs had a complete open reading frame (ORF) and encoded 78, 77 and 76 amino acid residues, respectively, including a strictly conserved γ-thionin domain of eight cysteines which is characteristic in plant defensins. Multiple alignment analysis based on the deduced amino acid sequences of defensin genes from different pIants indicated that NtDEF1, NtDEF2 and NtDEF3 showed high similarity with 50%-87% identities to the defensins from Capsicum annuum. However, with 33%-40% identities to the defensins previously reported from Nicotiana, the evolutionary analysis showed that NtDEF1, NtDEF2 and NtDEF3 were very far related to them. In conclusion, this is the first time that the NtDEF1, NtDEF2 and NtDEF3 genes have been reported in tobacco.
- Nicotiana tabacum /
- defensin /
- cloning /
- sequence analysis

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参考文献(19)

[1]	Broekaert W F, Terras F R, Cammue B P, et al. Plant defensins: novel antimicrobial peptides as components of the host defense system [J]. Plant Physiol, 1995, 108: 1353-1358.
[2]	Lai F M, DeLong C, Mei K, et al. Analysis of the DRR230 family of pea defensins: gene expression pattern and evidence of broad host-range antifungal activity [J]. Plant Sci, 2002, 163: 855-864.
[3]	Solis J, Medrano G, Ghislain M. Inhibitory effect of a defensin gene from the Andean crop maca (Lepidium meyenii) against Phytophthora infestans [J]. J Plant Physiol, 2007, 164: 1071-1082.
[4]	Choi Y, Choi Y D, Lee J S. Antimicrobial activity of γ-thionin-like soybean SE60 in E. coli and tobacco plants [J]. Biochem Biophys Res Commun, 2008, 375: 230-234.
[5]	Wijaya R, Neumann G M, Condron R, et al. Defense proteins from seed of Cassia fistula include a lipid transfer protein homologue and a protease inhibitory plant defensin [J]. Plant Sci, 2000, 159: 243-255.
[6]	Chen K C, Lin C Y, Kuan C C, et al. A novel defensin encoded by a mungbean cDNA exhibits insecticidal activity against bruchid [J]. J Agric Food Chem, 2002, 50: 7258-7263.
[7]	Do H M, Lee S C, Jung H W, et al. Differential expression and in situ localization of a pepper defensin (CADEF1) gene in response to pathogen infection, abiotic elicitors and environmental stresses in Capsicum annuum [J]. Plant Sci, 2004, 166: 1297-1305.
[8]	Gu Q, Kawata E E, Morse M J, et al. A flower-specific cDNA encoding a novel thionin in tobacco [J]. Mol Gen Genet, 1992, 234: 89-96.
[9]	Yamada S, Komori T, Imaseki H. cDNA cloning of γ-thionin from Nicotiana excelsior (Accession No. AB005266) (PGR97-131) [J]. Plant Physiol, 1997, 115: 314.
[10]	Lou Y, Baldwin I T. Nitrogen supply influences herbivore-induced direct and indirect defenses and transcriptional responses in Nicotiana attenuate [J]. Plant Physiol, 2004, 135: 496-506.
[11]	Lay F T, Brugliera F, Anderson M A. Isolation and properties of floral defensins from ornamental tobacco and petunia [J]. Plant Physiol, 2003, 131: 1283-1293.
[12]	Lay F T, Schirra H J, Scanlon M J, et al. The three-dimensional solution of NaD1, a new floral defensin from Nicotiana alata and its application to a homology model of the crop defense protein alfAFP [J]. J Mol Biol, 2003, 325: 175-188.
[13]	Gill R W, Sanseau P. Rapid in silico cloning of genes using expressed sequence tags (ESTs) [J]. Biotechnol Annu Rev, 2000, 5: 25-44.
[14]	Carvalho A O, Gomes V M. Plant defensins-prospects for the biological functions and biotechnological properties [J]. Peptides, 2009, 30: 1007-1020.
[15]	Pelegrini P B, Franco O L. Plant γ-thionins: novel insights on the mechanisms of actions of a multi-functional class of defense proteins [J]. Int J Biochem Cell Biol, 2005, 37: 2239-2253.
[16]	Mendez E, Moreno A, Colilla F, et al. Primary structure and inhibition of protein synthesis in eukaryotic cell-free system of a novel thionin, γ-hordothionin, from barley endosperm [J]. Eur J Biochem, 1990, 194: 533-539.
[17]	Bruix M, Jimenez M A, Santoro J, et al. Solution structure of gamma 1-H and gamma 1-P thionins from barley and wheat endosperm determined by 1H-NMR: a structural motif common to toxic arthropod proteins [J]. Biochemistry, 1993, 32: 715-724.
[18]	Colilla F J, Rocher A, Mendez E. γ-Purothionins: amino acid sequence of two polypeptides of a new family of thionins from wheat endosperm [J]. FEBS Lett, 1990, 270: 191-194.
[19]	Meyer B, Houlne G, Pozueta-Romero J, et al. Fruit-specific expression of a defensin-type gene family in bell pepper. Upregulation during ripening and upon wounding [J]. Plant Physiol, 1996, 112: 615-622.