A new bacteriophage P1–derived vector for the propagation of large human DNA fragments

Panayiotis A. Loannou, Lawrence Livermore National Laboratory
Chris T. Amemiya, Lawrence Livermore National Laboratory
Jeffrey Garnes, Lawrence Livermore National Laboratory
Peter M. Kroisel, Lawrence Livermore National Laboratory
Hiroaki Shizuya, California Institute of Technology
Chira Chen, Lawrence Livermore National Laboratory
Mark A. Batzer, Lawrence Livermore National Laboratory
Pieter J. de Jong, Lawrence Livermore National Laboratory

Abstract

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. © 1994 Nature Publishing Group.