Hamada H
Mol Cell Biol. 1986 Dec;6(12):4179-84. doi: 10.1128/mcb.6.12.4179-4184.1986.
Expression of enhancerless (E-) and enhancer-containing (E+) genes that are chromosomally integrated was examined. An E- plasmid (pE-cat) containing a chloramphenicol acetyltransferase (cat) gene linked to the simian virus 40 (SV40) early promoter or its E+ counterpart plasmid (pE+-cat) containing the SV40 enhancer was cotransfected into thymidine kinase (TK)-deficient L cells with a cloned tk gene. A number of TK+ transformants were isolated, and expression of the cointegrated cat gene in these cell lines was quantitatively determined by the assay of CAT activity. The results indicated unexpectedly that the E- cat gene was as actively expressed as the E+ cat gene. Analysis of CAT mRNA by primer extension indicated that the E- cat gene, as well as the E+ cat gene, was transcribed from the "native" initiation site contained in the SV40 early promoter region. The active expression of the E- cat gene was maintained in secondary TK+ transformants that arose by transfection with genomic DNA from the primary transformant. These results suggest that expression of the integrated E- cat gene is activated by endogenous enhancer elements.
对染色体整合的无增强子（E-）基因和含增强子（E+）基因的表达进行了检测。将含有与猴病毒40（SV40）早期启动子相连的氯霉素乙酰转移酶（cat）基因的E-质粒（pE-cat）或含有SV40增强子的其E+对应质粒（pE+-cat）与克隆的tk基因共转染到胸苷激酶（TK）缺陷的L细胞中。分离出许多TK+转化体，并通过检测CAT活性定量测定这些细胞系中共同整合的cat基因的表达。结果出乎意料地表明，E- cat基因与E+ cat基因一样活跃表达。通过引物延伸对CAT mRNA进行分析表明，E- cat基因以及E+ cat基因均从SV40早期启动子区域所含的“天然”起始位点转录。E- cat基因的活跃表达在通过用来自初级转化体的基因组DNA转染产生的次级TK+转化体中得以维持。这些结果表明，整合的E- cat基因的表达被内源性增强子元件激活。
