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A modification vector has been constructed to facilitate the transfer of yeast artificial chromosomes (YACs) to mammalian cells in culture by targeting a dominant selectable marker (G418 resistance) to the right arm of pYAC4 clones. The ADE2 gene is used for yeast selection with consequent disruption of the URA3 gene, allowing direct modification of YACs within the common host strain AB1380, and providing a simple test for correct targeting. This vector has been tested by modification of a 550-kb YAC containing part of the human MHC class II region and transfer to CHO cells by protoplast fusion. Analysis of 15 independent G418-resistant CHO lines obtained following fusion suggests the majority contain a complete YAC with moderate amplification in some lines.

Type

Journal article

Journal

Somat cell mol genet

Publication Date

03/1993

Volume

19

Pages

161 - 169

Keywords

Animals, Blotting, Southern, CHO Cells, Chromosomes, Fungal, Cricetinae, Drug Resistance, Gene Library, Genetic Vectors, Gentamicins, In Situ Hybridization, Fluorescence, Mammals, Transfection