Tzall S, Martiniuk F
New York University Medical Center, Department of Medicine, NY 10016.
Biochem Biophys Res Commun. 1991 May 15;176(3):1509-15. doi: 10.1016/0006-291x(91)90458-j.
Genetic deficiency of acid alpha glucosidase (GAA) results in glycogen storage disease type II. A cDNA containing the complete coding region was constructed and cloned into the expression vector pSV2 and was transiently transfected into an SV40 immortalized GAA deficient human fibroblast cell line which has undetectable levels of GAA enzyme activity and does not express GAA mRNA. Transfected cells had 4.9% of normal human fibroblast enzyme activity. Additionally a 5' 1.8 kb genomic fragment was ligated to the 5' end of the GAA cDNA construct and cloned into pUC19. Transient and stable transfection also resulted in expressed GAA enzyme activity in deficient fibroblast cells, indicating that the genomic fragment has GAA promoter function.
酸性α-葡萄糖苷酶(GAA)的基因缺陷会导致II型糖原贮积病。构建了一个包含完整编码区的cDNA,并将其克隆到表达载体pSV2中,然后瞬时转染到一种SV40永生化的GAA缺陷型人成纤维细胞系中,该细胞系的GAA酶活性水平检测不到,且不表达GAA mRNA。转染后的细胞具有正常人成纤维细胞4.9%的酶活性。此外,将一个5' 1.8 kb的基因组片段连接到GAA cDNA构建体的5'端,并克隆到pUC19中。瞬时转染和稳定转染也导致缺陷型成纤维细胞中表达GAA酶活性,表明该基因组片段具有GAA启动子功能。