HUMAN ARTIFICIAL EPISOMAL CHROMOSOMES FOR CLONING LARGE DNA FRAGMENTSIN HUMAN-CELLS

We have developed a human artificial episomal chromosome (HAEC) system , based on the latent replication origin of the large herpes Epstein-B arr virus, for the propagation and stable maintenance of DNA as circul ar minichromosomes in human cells. Individual HAECs carried human geno mic inserts ranging from 60-330 kb and appeared genetically stable. An HAEC library of 1,500 independent clones carrying random human genomi c fragments with average sizes of 150-200 kb was established and allow ed recovery of the HAEC DNA. Our autologous HAEC system, with human DN A cloned directly in human cells, provides an important tool for funct ional study of large mammalian DNA regions and gene therapy.