杉木纤维素合成酶基因CesA的克隆及表达分析

Isolation and expression analysis of cellulose synthase genes in Chinese fir(Cunninghamia lanceolata)

通过逆转录聚合酶链式反应（RT-PCR）结合c DNA末端快速扩增（RACE）技术,从杉木Cunninghamia lanceolata中克隆到2个全长为3 235 bp和3 876 bp的纤维素合成酶基因c DNA序列,被分别命名为Cl Ces A1和Cl Ces A2,Gen Bank登录号分别为JQ844574和JQ844575。相应编码蛋白分别包含992和1 092个氨基酸残基,推测分子量为111 845.3 D和123 105.7 D,等电点为6.04和6.65。氨基酸序列分析发现,它们具有植物纤维素合成酶基因相应的结构特征——锌指结构,2个易变区CSRI和CSRII,2个保守区CRI和CRII,纤维素合成酶底物结合结构域＂D,D,D,QVLRW＂,以及8个跨膜区。荧光定量PCR分析表明：Cl Ces A1和Cl Ces A2基因在茎中表达丰度均为最高,且成熟木质部中表达量均高于皮层中的相应数值;2个基因在根和针叶中的表达量相对较低。以具特殊材质性状的矮生杉木和正常杉木木质部为材料的进一步表达分析发现：Cl Ces A1和Cl Ces A2在正常杉木木质部中的表达量大约是矮生杉木表达量的2~12倍。2个杉木Ces A基因可能在杉木木材形成过程中具有重要作用。

In this study, two full-length cDNAs encoding cellulose synthase （CESA） were isolated from Chinese fir （Cunninghamia lanceolata） using RT-PCR and RACE technology, and their sequence lengths were 3 235 bp and 3 876 bp, respectively. They were named C1CesA 1 and C1CesA2, and the corresponding GenBank accession numbers were JQ844574 and JQ844575 respectively. The encoded proteins are composed of 995 and 1 092 amino acids, with theoretical molecular weight of 111 845.3 and 123 105.7 D, whose isoelectric points are 6.04 and 6.65, respectively. Amino acid sequences contain typical motifs of plant CESA proteins： a ring finger domain, two class specific regions CSRI and CSRII, two conserved regions CRI and CRII, cellulose synthase substrate binding domain ＂D, D, D, QVLRW＂ , and eight transmembrane regions. The expression analysis showed that ClCesA 1 and ClCesA2 genes exhibited the high transcript abundance in stem, the corresponding expression levels in mature xylem were higher than that in cortex. The two genes had the lower expression levels in root and leaf. The further expression analysis was conducted using the dwarf Chinese fir with the specific wood properties. The results showed that the expression levels of ClCesA 1 and CICesA 2 in the xylem of normal fir were about 2-12 times than that of the dwarf. Two ClCesA genes may be play important roles in the wood formation of Chinese fir.