体外促进血管内皮细胞分化的实验观察:人血管生成素相关蛋白2全长编码基因的克隆

Experimental observation on the differentiation of vascular endothelial cells facilitated in vitro:gene clone of angiopoietin-related protein 2

目的:克隆人血管生成素相关蛋白2(angiopoietin-relatedprotein2,ARP2)全长编码基因,使其以体外方式促进血管内皮细胞分化,拟构建PET32a载体。方法:采用反转录多聚酶链反应(RT-PCR)从人脾脏组织中扩增ARP2cDNA,构建PET32a载体,采用EcoRI,HindIII双酶切以及DNA测序进行鉴定。结果:RT-PCR扩增长度为1492bp的基因片段。EcoRI,HindIII双酶切结果显示重组T载体中含有目的基因片段,测序结果显示编码区无基因突变。结论:顺利构建PET32a载体,人ARP2全长cDNA基因克隆成功。ARP2有非常明显的促进血管内皮细胞生长的作用,可能对缺血性心脏病的促血管形成治疗有较大帮助。

AIM:To clone the full-length cDNA of human angiopoietin-related protein 2(ARP2) gene so as to facilitate the differentiation of vascular endothelial cells(VECs) in vitro,and construct PET32a vector. METHODS:The full-length cDNA of human APR2 was amplified successfully from human spleen tissues using RT-PCR technique,and then cloned into PET32a vector,which was identified and confirmed by restriction endonuclease mapping using EcoR I and Hind III as well as DNA sequence analysis. RESULTS:The human ARP2 DNA containing 1 492 bp was amplified using RT-PCR,restriction endonuclease mapping using EcoR I and Hind III showed that targeting DNA was inserted into recombinant T vector,and there was no mutation in the coding area through DNA sequencing. CONCLUSION:It was successful to construct PET32a vector.The human full-length ARP2 cDNA gene was successfully cloned.ARP2 has a significant role in facilitating the VECs differentiation,and it may be of great benefit to improving the vascularization for treatment of ischemic heart diseases.