Fountain J W, Lockwood W K, Collins F S
Howard Hughes Medical Institute, University of Michigan, Ann Arbor 48109.
Gene. 1988 Aug 15;68(1):167-72. doi: 10.1016/0378-1119(88)90610-5.
Primary human skin fibroblasts are an accessible source of phenotypically and karyotypically normal human cells, but are difficult to transfect with exogenous DNA. Here we demonstrate that both transient expression and stable transformation can be carried out by the method of electroporation. Highly efficient transient chloramphenicol acetyltransferase expression was shown after transfection with plasmid pRSVCAT. Stable transformation of human skin fibroblasts to G418 resistance was obtained after electroporation with neo-containing plasmids at an efficiency of approximately 1.4 x 10(-5)/micrograms DNA. The ability to easily transfect these cells with exogenous DNA may have important applications in the study of human genetic diseases and cancer.
原代人皮肤成纤维细胞是表型和核型正常的人类细胞的一个可获取来源,但用外源DNA转染它们很困难。在此我们证明,通过电穿孔法可以实现瞬时表达和稳定转化。用质粒pRSVCAT转染后显示出高效的瞬时氯霉素乙酰转移酶表达。用含neo的质粒进行电穿孔后,获得了对G418耐药的人皮肤成纤维细胞的稳定转化,效率约为1.4×10⁻⁵/微克DNA。用外源DNA轻松转染这些细胞的能力可能在人类遗传疾病和癌症研究中有重要应用。