《植物生理学报》 2012, 48(5): 449-455
通信作者:常有宏;E-mail: cyh@jaas.ac.cn;Tel: 025-84390224
摘 要:
采用RT-PCR、cDNA末端快速扩增法和长片段PCR技术, 从杜梨幼苗中获得1个磷酸乙醇胺N-甲基转移酶基因(Pb- PEAMT), 运用生物信息学方法分析它的序列特点, 并通过跨内含子引物进行半定量RT-PCR研究其在非生物胁迫下的表达 情况。结果表明: PbPEAMT基因编码区DNA序列长为3 320 bp, 由11个外显子和10个内含子组成, cDNA序列长1 479 bp, 推 导的多肽包括2个II型甲基转移酶保守结构域, 与蓖麻PEAMT蛋白相似性最高(86%), 亲缘关系最近。PbPEAMT基因在杜 梨幼苗根和叶中均为诱导型表达, 100 mmol•L-1氯化钠、10% (W/V)聚乙二醇、180 mmol•L-1甘露醇或20 µmol•L-1脱落酸处理后 PbPEAMT表达水平上升, 表明PbPEAMT对盐碱、干旱和渗透胁迫存在表达响应, 可能参与ABA介导的逆境信号转导途径。关键词:杜梨; 磷酸乙醇胺N-甲基转移酶; 基因克隆; 表达特征
收稿:2011-12-14 修定:2012-02-29
资助:国家自然科学基金(31101529)和江苏省农业科技自主创新 资金[CX(11)4050]。
Corresponding author: CHANG You-Hong; E-mail: cyh@jaas.ac.cn; Tel: 025-84390224
Abstract:
N-methylation of phosphoethanolamine, the committing step in choline biosynthesis in plants, is catalyzed by phosphoethanolamine N-methyltransferase (PEAMT, EC 2.1.1.103). Herein we report the cloning and characterization of a novel birch-leaf pear (Pyrus betulaefolia) phosphoethanolamine N-methyltransferase gene (PbPEAMT) using a combination of homologous cloning, PCR and bioinformatics strategy. At the same time, semi-quantitative PCR with cross-intron primers was adopted to study the expression features of this gene under abiotic stresses. The results showed that the DNA sequence of PbPEAMT gene is 3 320 bp, which consists of 11 exons and 10 introns. The cDNA sequence of PbPEAMT gene is 1 479 bp in length and encodes a 492-amino-acid peptide, which includes two conserved motifs of type II methyltransferase. Homology analysis showed that the deduced PbPEAMT protein was highest homologous to castor PEAMT protein (86%). Furthermore, PbPEAMT was more related to castor PEAMT through phylogenetic analysis. RT-PCR analysis showed that expression of PbPEAMT was induced by 100 mmol·L-1 NaCl, 10% (W/V) PEG6000, 180 mmol·L-1 mannitol or 20 µmol·L-1ABA treatments. These data indicate that PbPEAMT expression responds to salinity, drought and osmotic stresses, which may be involved in ABA-mediated stress signal transduction pathway.Key words: birch-leaf pear (Pyrus betulaefolia); phosphoethanolamine N-methyltransferase; gene cloning; expression feature
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