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PMID: 1704929 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Retrovirus promoter-trap vector to induce lacZ gene fusions in mammalian cells.

Journal of virology ·Vol. 65 ·No. 3 ·1991-03-00 ·Pages 1507-15

Reddy S, DeGregori JV, von Melchner H, Ruley HE

Abstract

A retrovirus promoter-trap vector (U3LacZ) has been developed in which Escherichia coli lacZ coding sequences were inserted into the 3' long terminal repeat (LTR) of an enhancerless Moloney murine leukemia virus. The U3LacZ virus contains the longest reported LTR (3.4 kbp); nevertheless, lacZ sequences did not interfere with the ability of the virus to transduce a neomycin resistance gene expressed from an internal promoter. Duplication of the LTR placed lacZ sequences in the 5' LTR just 30 nucleotides from the flanking cellular DNA. Approximately 0.4% of integrated proviruses expressed beta-galactosidase as judged by 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside (X-Gal) staining, and individual clones expressing lacZ were isolated by fluorescence-activated cell sorting. In all clones examined, beta-galactosidase expression resulted from the fusion of lacZ sequences to transcriptional promoters located in the flanking cellular DNA. Furthermore, by differential sorting of neomycin-resistant cell populations, clones were isolated in which lacZ expression was induced and repressed in growth-arrested and log phase cells, respectively.

Related Genes
lacZ
MeSH Terms
Animals Blotting, Northern Blotting, Southern Cell Line Clone Cells Escherichia coli/enzymology,genetics Genetic Vectors Methionine/metabolism Mice Moloney murine leukemia virus/genetics Poly A/genetics,isolation & purification Promoter Regions, Genetic RNA/genetics,isolation & purification RNA, Messenger Recombinant Fusion Proteins/biosynthesis Repetitive Sequences, Nucleic Acid Restriction Mapping Transcription, Genetic Transfection beta-Galactosidase/biosynthesis,genetics
Chemicals
RNA, Messenger Recombinant Fusion Proteins Poly A RNA Methionine beta-Galactosidase
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Reddy S
Center for Cancer Research, Massachusetts Institute of Technology, Cambridge 02139.
DeGregori J V
von Melchner H
Ruley H E
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Article Info
Journal
Journal of virology
Abbr.
J Virol
ISSN
0022-538X
DOI
10.1128/JVI.65.3.1507-1515.1991
Published
1991-03-00
Pages
1507-15
Language
English
Region
United States
NLM ID
0113724
PMCID
PMC239931
Subset
IM
Grants
NCI NIH HHS · CA14051 · United States
NCI NIH HHS · P01CA42063 · United States
External Links
PubMed source DOI: 10.1128/JVI.65.3.1507-1515.1991 PMC Full Text View journal details
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