摘要
利用本地BLAST工具,从先期获得的苎麻转录组数据中发掘出与多种植物纤维素合成酶基因有较高同源性的序列CL6473。以苎麻(Boehmeria nivea)栽培种湘苎3号为材料,根据CL6473设计引物,先采用PCR扩增克隆了苎麻一个纤维素合成酶基因cDNA的核心片段,再运用5'及3'RACE获得该基因全长cDNA,序列分析表明该cDNA为一个新的苎麻纤维素合成酶基因cDNA,命名为BnCesA4。BnCesA4 cDNA序列全长4 008 bp,其中编码区全长3 270 bp,编码一个含1 090 aa的多肽,具有植物纤维素酶的保守结构域及特征氨基酸序列。根据该cDNA高可变区序列设计其特异性引物,采用实时荧光定量PCR(qRT-PCR)对BnCesA4基因在几个代表性苎麻品种:湘苎3号、湘苎1号、湘潭大叶白及城步青麻的木质部和韧皮部两种组织中的表达情况进行了定量分析。qRTPCR显示BnCesA4基因在不同品种苎麻的木质部及韧皮部均有表达,表达量差异不大。
The local BLAST comparison was run based on the ramie transcriptome data that previously obtained with cellu -lose synthase gene cDNA sequence of high plants as the probe ,and a potentially high homologous fragment CL 6473 was i-dentified from the database.A pairs of specific primers was designed according to CL 6473 to clone this fragments by core sequences PCR amplification and then followed with 3′RACE and 5′RACE from Boehmeria nivea variety Xiangzu No.3.A whole length cDNA was cloned that identity as a novel cellulose synthase gene cDNA ,named as BnCesA4.The full-length of BnCesA4 is 4 008 bp,and encodes a putative 1 090 amino acids protein.The cloned cDNA sequences were confirmed as ramie synthase genes by online BLAST and analysis of its conserved domains.The BnCesA4 expression in phloem and core stick tissues of ramie stems in four representative cultivars ,Xiangzu 3,Xiangzu 1,Xiangtaidayebai and Chengbuqingma were quantitated by qRT-PCR.The results showed that the BnCesA4 was both actively expressed in the phloem and core stick of ramie stems in the four different cultivars of ramie.The expression of BnCesA4 was in a closing level in the two or-gans as well.
出处
《作物研究》
2014年第5期472-478,共7页
Crop Research
基金
国家自然科学基金(31071457)
湖南省科技计划重点项目(2012NK3062)
湖南省教育厅项目(SCX1103)
湖南省教育厅科学研究一般项目(12C0156)
关键词
苎麻
纤维素合成酶基因
克隆
定量表达分析
Ramie
Cellulose synthase genes
Cloning
Quantitative expression analysis
