A shuttle system for transfer of YACs between yeast and mammalian cells | Zendy

Kyle A. Simpson | Zendy

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A shuttle system for transfer of YACs between yeast and mammalian cells

Author(s) -

Kyle A. Simpson

Publication year - 1996

Publication title -

nucleic acids research

Language(s) - English

Resource type - Journals

SCImago Journal Rank - 9.008

H-Index - 537

eISSN - 1362-4954

pISSN - 0305-1048

DOI - 10.1093/nar/24.23.4693

Subject(s) - biology , extrachromosomal dna , spheroplast , shuttle vector , autonomously replicating sequence , plasmid , yeast artificial chromosome , saccharomyces cerevisiae , dna , yeast , dna replication , transformation (genetics) , microbiology and biotechnology , origin of replication , genetics , recombinant dna , chromosome , gene , vector (molecular biology) , escherichia coli , gene mapping

The development of a system for shuttling DNA cloned as yeast artificial chromosomes (YACs) between yeast and mammalian cells requires that the DNA is maintained as extrachromosomal elements in both cell types. We have recently shown that circular YACs carrying the Epstein-Barr virus origin of plasmid replication (oriP) are maintained as stable, episomal elements in a human kidney cell line constitutively expressing the viral transactivator protein EBNA-1. Here, we demonstrate that a 90-kb episomal YAC can be isolated intact from human cells by a simple alkaline lysis procedure and shuttled back into Saccharomyces cerevisiae by spheroplast transformation. In addition, we demonstrate that the 90-kb YAC can be isolated intact from yeast cells. The ability to shuttle large, intact fragments of DNA between yeast and human cells should provide a powerful tool in the manipulation and analysis of functional regions of mammalian DNA.

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