Nature Genetics
6, 84 - 89 (1994)
doi:10.1038/ng0194-84
A new bacteriophage P1−derived vector for the propagation of large human DNA fragmentsPanayiotis A. loannou1, 4, Chris T. Amemiya1, 5, Jeffrey Garnes1, Peter M. Kroisel1, 6, Hiroaki Shizuya2, Chira Chen1, 3, Mark A. Batzer1
& Pieter J. de Jong1, 3, 7
1Human Genome Center L-452, Biology and Biotechnology Research Program, Lawrence Livermore National Laboratory, Livermore, California 94551, USA
2Division of Biology, 147-75, California Institute of Technology, Pasadena, California 91125, USA
3Human Genetics Department, Roswell Park Cancer Institute, Elm & Carlton Streets, Buffalo, New York 14263, USA
4Current address: The Cyprus Institute of Neurology and Genetics, P.O. Box 3462, Nicosia, Cyprus
5Current address: Center for Human Genetics, Boston University School of Medicine, 80 East Concord St, Boston, Massachusetts 02118, USA
6Current address: Department of Medical Biology and Human Genetics, Harrachgasse 21/8, A-8010, Graz, Austria
7Correspondence should be addressed to P.J.d.J. at Roswell Park Cancer Institute We have designed a P1 vector (pCYPAC−1) for the introduction of recombinant DNA into E. coli using electroporation procedures. The new cloning system, P1−derived arteficial chromosomes (PACs), was used to establish an initial 15,000 clone library with an average insert size of 130−150 kilobase pairs (kb). No chimaerism has been observed in 34 clones, by fluorescence in situ hybridization. Similarly, no insert instability has been observed after extended culturing, for 20 clones. We conclude that the PAC cloning system will be useful in the mapping and detailed analysis of complex genomes.
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