黑斑原鮡皮肤组织结构及其COL1A1基因的克隆与分析

Tissue Structure of Skin and Cloning and Analysis of COL1A1 Gene of the Glyptosternum Maculatum

采用组织学与分子生物学方法开展了黑斑原鮡皮肤组织结构和Ⅰ型胶原蛋白基因的表达特征研究。结果表明:(1)黑斑原鮡的皮肤裸露无鳞,均由表皮和真皮组成。黑斑原鮡表皮层由复层上皮细胞和黏液细胞以及基底细胞组成。其表皮层的厚度显著大于真皮层厚度,且不同部位表皮层的厚度各不同,但各部位真皮层的厚度接近。其中,腹部表皮层的厚度最厚,为(102.36±3.43)μm;头部最薄,为(66.91±9.73)μm。(2)黑斑原鮡头部、背部色素细胞含量丰富,腹部色素细胞含量较少。(3)黑斑原鮡Ⅰ型胶原蛋白的α1基因(COL1A1)的cDNA全长序列为6683 bp,其开放阅读框为4350 bp,编码1449个氨基酸。(4)BLAST同源性分析结果显示,黑斑原鮡COL1A1基因的氨基酸序列与巨魾的同源性最高,为92.06%,呈现出较高的保守性。(5)系统进化树分析表明,该基因与巨魾处于同一支,亲缘性最近。(6)生物信息学分析显示,黑斑原鮡COL1A1蛋白质的相对分子质量为137.14 KDa,理论等电点是5.42,为α螺旋β折叠三重螺旋结构蛋白,含有1个COLFI结构域、1个VWC结构域、1个RPT1结构域、2个RPT3结构域和8个Pfam Collagen结构域。

Histological and molecular biological methods were used to study the tissue structure of skin and ex⁃pression characteristics of typeⅠcollagen gene alpha 1in Glyptosternum maculatum.The result showed that(1)skin of the Glyptosternum maculatum was without scales and naked which was composed by epidermis and der⁃mis.The epidermis of the G.maculatum could be divided into stratified epithelium cell,goblet cell and basement membrane cell.Thicknesses of epidermis part was significantly higher than the dermis section,the abdominal part had highest thicknesses((102.36±3.43)μm),and the section of head took lowest thicknesses((66.91±9.73)μm).(2)Amount of pigment cell was higher at head and back part compare to abdominal part of the G.macula⁃tum.(3)The complete cDNA of typeⅠcollagen alpha 1(COL1A1)gene in G.maculatum consists of 6683 bp with an open reading frame of 4350 bp,encoding a protein of 1449 amino acids.(4)The result of BLAST homol⁃ogy analysis showed that the amino acids sequence of COL1A1 in G.maculatum had highest homology with the Bagarius yarrelli,and that was 92.06%,indicating higher conservatism.(5)The analysis of phylogenetic trees showed that this gene was belonging to the same branch with B.yarrelli,which indicated closest relationship be⁃tween them.(6)Bioinformatics analysis showed that relative molecular weight of COL1A1 protein in G.macula⁃tum was 137.14 KDa,and the theoretical isoelectric point was 5.42,which wasαhelix andβpleated sheet with three helix structure of protein,it involved 1 COLFI,1 VWC,1 RPT1,2 RPT3 and 8 Pfam Collagen structural do⁃mains.