VCAM-1基因的克隆及VCAM-1-pcDNA3.1(+)真核表达载体的构建

摘要

目的克隆人血管细胞黏附分子(vascular cell adhesion molecule-1,VCAM-1) cDNA全长,并构建VCAM-1-pcDNA3.1(+)真核表达载体.方法采用半巢式RT-PCR从人脐静脉内皮细胞系HUV-EC-C中分三段扩增得到VCAM-1cDNA片段,采用重叠延伸剪切技术(SOE)并经SacⅠ酶切、T4连接酶连接获得VCAM-1完整片段,克隆至pcDNA3.1(+)真核表达载体,得到VCAM-1-pcDNA3.1(+)真核表达载体.结果酶切及测序结果证明VCAM-1-pcDNA3.1(+)真核表达载体构建成功.结论成功扩增人VCAM-1基因,构建VCAM-1-pcDNA3.1(+)真核表达载体,为探索VCAM-1修饰人脐血基质细胞(human umbilical cord blood stromal cell,hUCBSC)重建造血微环境奠定了实验基础.