期刊
NUCLEIC ACIDS RESEARCH
卷 29, 期 3, 页码 -出版社
OXFORD UNIV PRESS
DOI: 10.1093/nar/29.3.e14
关键词
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资金
- Wellcome Trust Funding Source: Medline
Bacterial artificial chromosomes (BACs) and P1 artificial chromosomes (PACs), which contain large fragments of genomic DNA, have been successfully used as transgenes to create mouse models of dose-dependent diseases. They are also potentially valuable as transgenes for dominant diseases given that point mutations and/or small rearrangements can be accurately introduced. Here, we describe a new method to introduce small alterations in BACs, which results in the generation of point mutations with high frequency. The method involves homologous recombination between the original BAC and a shuttle vector providing the mutation. Each recombination step is monitored using positive and negative selection markers, which are the Kanamycin-resistance gene, the sacB gene and temperature-sensitive replication, all conferred by the shuttle plasmid. We have used this method to introduce four different point mutations and the insertion of the beta-galactosidase gene in a BAC, which has subsequently been used for transgenic animal production.
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