Gasser C S, Simonsen C C, Schilling J W, Schimke R T
Proc Natl Acad Sci U S A. 1982 Nov;79(21):6522-6. doi: 10.1073/pnas.79.21.6522.
We have constructed a number of abbreviated dihydrofolate reductase (DHFR) genes by using cloned mouse genomic and cDNA sequences. These genes contain 1.0 kilobase of 5' flanking genomic sequence and varying portions of the 3' non-coding region. Two of the genes contain the first two introns of the DHFR gene; the other three lack introns. Transfection of DHFR-deficient Chinese hamster ovary cells with any of these constructed genes results in cells with the DHFR+ phenotype. Treatment of the transfectants with methotrexate, a folate antagonist, leads to the emergence of methotrexate-resistant colonies which have amplified the transfected genes.
我们利用克隆的小鼠基因组和cDNA序列构建了多个简化的二氢叶酸还原酶(DHFR)基因。这些基因包含1.0千碱基的5'侧翼基因组序列和3'非编码区的不同部分。其中两个基因包含DHFR基因的前两个内含子;另外三个没有内含子。用这些构建的基因中的任何一个转染缺乏DHFR的中国仓鼠卵巢细胞,会产生具有DHFR+表型的细胞。用叶酸拮抗剂甲氨蝶呤处理转染细胞,会导致出现对甲氨蝶呤耐药的菌落,这些菌落扩增了转染的基因。