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PMID: 14704360 Published · epublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Facile methods for generating human somatic cell gene knockouts using recombinant adeno-associated viruses.

Nucleic acids research ·Vol. 32 ·No. 1 ·2004-01-02 ·Pages e3

Kohli M, Rago C, Lengauer C, Kinzler KW, Vogelstein B

Abstract

Emerging evidence suggests that recombinant adeno-associated viral (rAAV) vectors can be used for specific gene targeting in human somatic cells. We have developed an rAAV vector construction procedure employing fusion PCR and a single cloning step that considerably simplifies the knockout process. We demonstrate its utility by disrupting genes at specific positions within human colon cancer cells as well as within immortalized normal epithelial cells. This technology should be broadly applicable to in vitro studies that require the manipulation of the human genome.

MeSH Terms
Cell Line Cell Line, Tumor Colonic Neoplasms/metabolism Dependovirus/genetics Epithelial Cells/metabolism Gene Deletion Gene Targeting/methods Genetic Vectors/genetics Humans Mutagenesis, Site-Directed Plasmids/genetics Polymerase Chain Reaction
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Kohli Manu
The Howard Hughes Medical Institute, The Sidney Kimmel Comprehensive Cancer Center, and The Cellular and Molecular Medicine Program, The Johns Hopkins University Medical Institutions, Baltimore, MD 21231, USA.
Rago Carlo
Lengauer Christoph
Kinzler Kenneth W
Vogelstein Bert
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Article Info
Journal
Nucleic acids research
Abbr.
Nucleic Acids Res
ISSN
1362-4962
DOI
10.1093/nar/gnh009
Published
2004-01-02
Epub
2004-00-02
Pages
e3
Language
English
Region
England
NLM ID
0411011
PMCID
PMC373311
Subset
IM
Grants
NCI NIH HHS · CA 57345 · United States
NCI NIH HHS · R37 CA043460 · United States
NCI NIH HHS · P50 CA062924 · United States
NCI NIH HHS · CA 43460 · United States
NCI NIH HHS · CA 62924 · United States
NCI NIH HHS · R37 CA057345 · United States
NCI NIH HHS · R01 CA057345 · United States
External Links
PubMed source DOI: 10.1093/nar/gnh009 PMC Full Text View journal details
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