Retroviral insertional mutagenesis of a target allele in a heterozygous murine cell line

W. Frankel, T. A. Potter, N. Rosenberg, Jack Lenz, T. V. Rajan

Research output: Contribution to journalArticle

18 Citations (Scopus)

Abstract

A clonal murine cell line that is heterozygous at the β2-microglobulin locus (B2m) was obtained by Abelson murine leukemia virus (Ab-MuLV) transformation of liver cells from (C57BL/6 x BALB/c) F1 fetuses. To obtain proviral insertional mutants, we superinfected a subclone of these cells, which does not express the env surface protein of the Moloney leukemia virus (Mo-MuLV, the helper virus that was used to transmit the defective Ab-MuLV genome during transformation), with Mo-MuLV. Mutant clones that fail to express the C57BL/6 allele of B2m (B2mb) were then immunoselected by using a monoclonal antibody that specifically recognizes the B2mb gene product and not that of the B2ma allele. Of 22 independent clones obtained, one contains a proviral insertion that is near or in the first exon of the B2mb gene. Surprisingly, the insertion is of the Ab-MuLV genome and not of replication-competent Mo-MuLV. This indicates that superinfection with Mo-MuLV 'rescued' the defective Ab-MuLV genome, which then inserted into the B2mb gene. We conclude that when an allele-specific selection procedure exists, proviral insertion is a potential method for obtaining mutations in heterozygous mammmalian cells. This approach may thereby provide a method for molecular cloning of such selectable genes, using a retroviral hybridization probe.

Original languageEnglish (US)
Pages (from-to)6600-6604
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume82
Issue number19
DOIs
StatePublished - 1985
Externally publishedYes

Fingerprint

Abelson murine leukemia virus
Insertional Mutagenesis
Alleles
Defective Viruses
Cell Line
Genome
Genes
Clone Cells
Helper Viruses
env Gene Products
Moloney murine leukemia virus
Superinfection
Molecular Cloning
Exons
Membrane Proteins
Fetus
Monoclonal Antibodies
Mutation
Liver

ASJC Scopus subject areas

  • Genetics
  • General

Cite this

Retroviral insertional mutagenesis of a target allele in a heterozygous murine cell line. / Frankel, W.; Potter, T. A.; Rosenberg, N.; Lenz, Jack; Rajan, T. V.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 82, No. 19, 1985, p. 6600-6604.

Research output: Contribution to journalArticle

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AU - Potter, T. A.

AU - Rosenberg, N.

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AU - Rajan, T. V.

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