毛竹纤维素合成酶基因PeCesA的克隆及组织表达谱分析

Cloning and Expression Characterization of the Cellulose Synthase Gene (PeCesA) from Moso Bamboo (Phyllostachys edulis) Shoot

根据物种间同源基因设计引物,对构建好的毛竹笋全长cDNA文库进行大规模PCR筛选,成功分离出了毛竹纤维素合成酶基因PeCesA的cDNA序列,全长共3545bp(GenBank登录号:FJ799713)。该序列包含3243bp的开放阅读框(ORF),编码1081个氨基酸。通过核酸序列比对发现,该序列与绿竹、水稻、玉米、小麦和大麦的纤维素合成酶基因同源性极高(>90%)。蛋白序列比对分析发现:PeCesA蛋白含有植物纤维素合成酶特有的保守区、可变区、N端Zn指结构域以及8个跨膜区;系统发育分析表明PeCesA与绿竹BoCesA4、水稻OsCesA4、玉米ZmCesA4属于同一进化分支。运用实时定量PCR法分析PeCesA基因的组织表达特异性发现,PeCesA在毛竹根部的表达量最高,茎部其次,叶中较少;在竹笋基部的表达量远远高于竹笋上部。随着PeCesA基因表达量的增高,组织中的纤维素含量亦相应增高。推测PeCesA参与了毛竹次生细胞壁中纤维素的生物合成。

The cellulose synthase gene（CesA）plays a key role in regulating cellulose biosynthesis.A 3 545 bp cDNA clone encoding PeCesA（GenBank accession No.FJ799713）was isolated from the full-length cDNA library of the shoot of Moso bamboo（Phyllostachys edulis）by PCR screen.The cDNA had an 3 243 bp open reading frame（ORF）which would be capable to encode 1 081 amino acid residues.Multiple alignment analyses showed that the PeCesA shared high similarity（over 90%）with the CesA genes from Bambusa oldhamii,Oryza sativa,Triticum aestivum and Hordeum vulgare.And the deduced amino acid sequence of PeCesA contained highly conserved two Zinc-binding domains , eight transmembrance domains and one QVLRW motif,which were special characteristics among the cellulose synthase of plant.Phylogenetic analysis showed that PeCesA was clustered in the group that included CesA4s from Bambusa oldhamii,Oryza sativa and Zea mays respectively.The expression profile of PeCesA was analyzed by quantitative Real Time PCR.The results showed that the expression of PeCesA in the top of the bamboo shoots was extremely low,but it was high in the base of shoot.And it was relatively lower in the leaves compared to the roots and stems.The results of determination of cellulose content showed there was correlation between the expression of PeCesA and the cellulose content in different organisms.It suggested that PeCesA involved in cellulose synthesis in second cell wall Moso bamboo shoots.