黑麦草漆酶编码基因片段的克隆与序列分析

黑麦草中木质素的结构、含量是决定其饲用品质的主要因素。漆酶是木质素生物合成中的关键酶之一,属于多铜氧化酶基因家族。根据报道的植物漆酶编码基因的氨基酸序列中铜离子结合域及N-端糖基化位点两个保守区设计4对引物,以黑麦草总DNA为模板,经PCR扩增、克隆、测序后得到3个有效序列,分别命名为Lac10-1、Lac8-1和Lac11-1。序列分析表明,3个基因片段的编码蛋白不仅含有保守的蓝铜氧化酶结合区域HAH和N-末端糖基化位点,而且分别与黄杨、黑麦草中分离的漆酶编码蛋白的氨基酸序列具有较高的同源性。因而,推断克隆到的3个基因片段是黑麦草漆酶基因家族的成员。

Cloning and sequence analysis of laccase -coding gene fragments from ryegrass (Lolium perenne )

Forage quality of ryegrass(Lolium perenne) is mainly affected by the structure,content of lignin.Plant laccases belonged to the large multicopper oxidase gene family,are believed to be involved in lignin biosynehsis.Primers used in this work were designed according to the conservative amino acid regions in laccase protein family.PCR amplifications were performed on ryegrass(Lolium perenne) genomic DNA template.Three different laccase-encoding gene fragments,Lac10-1,Lac8-1 and Lac11-1 were identified finally.By using the program of BLAST on NCBI GenBank database,the sequences presented high matches with the laccase genes from other plants.The results indicated that the cloned sequences with conserved blue copper oxidase signatures and potential N-glycosylation sites shared higher homology at amino acid level to laccase-encoding cDNA sequences,LtLACC2-3 from Liriodendron tulipifera,LpLAC11 and LpLAC56 from Lolium perenne.So the cloned fragments belong to the members of laccase-coding gene fragment from ryegrass were concluded.