Riley J H, Morten J E, Anand R
ICI Pharmaceuticals, Biotechnology Department, Macclesfield, Cheshire, UK.
Nucleic Acids Res. 1992 Jun 25;20(12):2971-6. doi: 10.1093/nar/20.12.2971.
Vectors have been constructed for the introduction of the neomycin resistance gene (neo) into the left arm, right arm or human insert DNA of yeast artificial chromosomes (YACs) by homologous recombination. These vectors contain a yeast selectable marker Lys-2, i.e. the alpha-aminoadipidate reductase gene, and a mammalian selection marker, neo, which confers G418 resistance. The vectors can be used to modify YACs in the most commonly used yeast strain for YAC library construction, AB1380. Specific targeting can be carried out by transfection of restriction endonuclease treated linear plasmids, with highly specific recombinogenic ends, into the YAC containing yeast cells. Analysis of targeted YACs confirmed that all three vectors can target correctly in yeast. Introduction of one of the targeted YACs into V79 (Chinese hamster fibroblast) cells showed complete and intact transfer of the YAC.
已经构建了载体，用于通过同源重组将新霉素抗性基因（neo）引入酵母人工染色体（YAC）的左臂、右臂或人类插入DNA中。这些载体包含酵母选择标记Lys-2，即α-氨基己二酸还原酶基因，以及赋予G418抗性的哺乳动物选择标记neo。这些载体可用于在构建YAC文库最常用的酵母菌株AB1380中修饰YAC。通过将经限制内切酶处理的具有高度特异性重组末端的线性质粒转染到含有YAC的酵母细胞中，可以进行特异性靶向。对靶向YAC的分析证实，所有三种载体都能在酵母中正确靶向。将其中一个靶向YAC引入V79（中国仓鼠成纤维细胞）细胞，显示YAC完全完整地转移。
