中国农业科学2012，Vol.45Issue(1)：118-126,9.DOI:10.3864/j.issn.0578-1752.2012.01.014
龙眼咖啡酰辅酶A-O-甲基转移酶(DLCCoAOMT)基因的克隆和表达分析
Molecular Cloning of Longan Caffeoy(I)-CoA O-methyltransferase (DLCCoAOMT) and Its Expression Analysis
摘要
Abstract
[Objective] The aim of this study was to clone full-length cDNA of a key enzyme gene DLCCoAOMT related to lignin metabolism in longan, investigate its sequence characteristics and analyze its expression in different organs under low temperature stress, as well as expression in the prokaryote. [Method] The DLCCoAOMT gene cDNA sequence was cloned from longan leaf using RT-PCR and RACE techniques. The bioinformatics methods were used to analyze putative amino acid sequence and real-time PCR method were used to study the expression of DLCCoAOMT gene in different tissues. [ Result ] The full-length cDNA of DLCCoAOMT (GenBank accession number: JN093023) in longan was cloned. The sequence consists of 993 bp with an open reading frame of 744 bp, encoding a polypeptide of 247 amino acids. Homology analysis showed that the deduced DLCCoAOMT protein was highly homologous to other CCoAOMT proteins from different species. Phylogenetic analysis also indicated that DLCCoAOMT was very closely related to DLCCoAOMT of Betula. Real-time PCR results showed that the DLCCoAOMT expressed in root, stem and leaf, and its expression was different among three organs. The mRNA of DLCCoAOMT was abundant in root and stem, but less in leaf. Furthermore, DLCCoAOMT transcription level was significantly different among root, stem and leaf with time courses of chilling treatment. Prokaryotic expression showed that recombinant plasmid was efficiently expressed in Escherichia coli BL21. [ Conclusion ] Gene DLCCoA OMT was firstly isolated and characterized from longan, which may be involved in chilling stress.关键词
龙眼/咖啡酰辅酶A-O-甲基转移酶/基因克隆/表达分析/原核表达Key words
Dimocarpus longan Lour./caffeoyl CoA 3-O-methyltransferase/gene cloning/expression analysis/prokaryotic expression引用本文复制引用
陈虎,何新华,罗聪,杨丽涛,张保青,宋修鹏..龙眼咖啡酰辅酶A-O-甲基转移酶(DLCCoAOMT)基因的克隆和表达分析[J].中国农业科学,2012,45(1):118-126,9.基金项目
国家科技支撑计划项目(2008BADB8B01,2008BADB8B02)、广西自然科学基金项目(2011GXNSFA018115)、广西高等学校优秀人才计划项目(桂教人201065)、广西研究生创新计划项目(GXU11T31077) （2008BADB8B01,2008BADB8B02）
