Efficiency of insertion versus replacement vector targeting varies at different chromosomal loci

Paul Hasty, Mike Crist, Markus Grompe, Allan Bradley

Research output: Contribution to journalArticle

30 Citations (Scopus)

Abstract

We have analyzed the targeting frequencies and recombination products generated with isogenic vectors at the fah and fgr loci in embryonic stem cells. A single vector which could be linearized at different sites to generate either a replacement or an insertion vector was constructed for each locus. A replacement event predominated when the vectors were linearized at the edge of the homologous sequences, while an insertion event predominated when the vectors were linearized within the homologous sequences. However, the ratio of the targeting frequencies exhibited by the different vector configurations differed for the two loci. When the fgr vector was linearized as an insertion vector, the ratio of targeted to random integrations was four- to eightfold greater than when the vector was linearized as a replacement vector. By contrast, the ratio of targeted to random integrations at the fah locus did not vary with the linearization site of the vector. The different relationships between the targeting frequency and the vector configuration at the fgr and fah loci may indicate a DNA sequence or chromatin structure preference for different targeting pathways.

Original languageEnglish (US)
Pages (from-to)8385-8390
Number of pages6
JournalMolecular and Cellular Biology
Volume14
Issue number12
StatePublished - Dec 1994

Fingerprint

Sequence Homology
Insertional Mutagenesis
Embryonic Stem Cells
Genetic Recombination
Chromatin

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics
  • Cell Biology

Cite this

Efficiency of insertion versus replacement vector targeting varies at different chromosomal loci. / Hasty, Paul; Crist, Mike; Grompe, Markus; Bradley, Allan.

In: Molecular and Cellular Biology, Vol. 14, No. 12, 12.1994, p. 8385-8390.

Research output: Contribution to journalArticle

@article{6bc96cd22cfd42d687b5e303e54ad390,
title = "Efficiency of insertion versus replacement vector targeting varies at different chromosomal loci",
abstract = "We have analyzed the targeting frequencies and recombination products generated with isogenic vectors at the fah and fgr loci in embryonic stem cells. A single vector which could be linearized at different sites to generate either a replacement or an insertion vector was constructed for each locus. A replacement event predominated when the vectors were linearized at the edge of the homologous sequences, while an insertion event predominated when the vectors were linearized within the homologous sequences. However, the ratio of the targeting frequencies exhibited by the different vector configurations differed for the two loci. When the fgr vector was linearized as an insertion vector, the ratio of targeted to random integrations was four- to eightfold greater than when the vector was linearized as a replacement vector. By contrast, the ratio of targeted to random integrations at the fah locus did not vary with the linearization site of the vector. The different relationships between the targeting frequency and the vector configuration at the fgr and fah loci may indicate a DNA sequence or chromatin structure preference for different targeting pathways.",
author = "Paul Hasty and Mike Crist and Markus Grompe and Allan Bradley",
year = "1994",
month = "12",
language = "English (US)",
volume = "14",
pages = "8385--8390",
journal = "Molecular and Cellular Biology",
issn = "0270-7306",
publisher = "American Society for Microbiology",
number = "12",

}

TY - JOUR

T1 - Efficiency of insertion versus replacement vector targeting varies at different chromosomal loci

AU - Hasty, Paul

AU - Crist, Mike

AU - Grompe, Markus

AU - Bradley, Allan

PY - 1994/12

Y1 - 1994/12

N2 - We have analyzed the targeting frequencies and recombination products generated with isogenic vectors at the fah and fgr loci in embryonic stem cells. A single vector which could be linearized at different sites to generate either a replacement or an insertion vector was constructed for each locus. A replacement event predominated when the vectors were linearized at the edge of the homologous sequences, while an insertion event predominated when the vectors were linearized within the homologous sequences. However, the ratio of the targeting frequencies exhibited by the different vector configurations differed for the two loci. When the fgr vector was linearized as an insertion vector, the ratio of targeted to random integrations was four- to eightfold greater than when the vector was linearized as a replacement vector. By contrast, the ratio of targeted to random integrations at the fah locus did not vary with the linearization site of the vector. The different relationships between the targeting frequency and the vector configuration at the fgr and fah loci may indicate a DNA sequence or chromatin structure preference for different targeting pathways.

AB - We have analyzed the targeting frequencies and recombination products generated with isogenic vectors at the fah and fgr loci in embryonic stem cells. A single vector which could be linearized at different sites to generate either a replacement or an insertion vector was constructed for each locus. A replacement event predominated when the vectors were linearized at the edge of the homologous sequences, while an insertion event predominated when the vectors were linearized within the homologous sequences. However, the ratio of the targeting frequencies exhibited by the different vector configurations differed for the two loci. When the fgr vector was linearized as an insertion vector, the ratio of targeted to random integrations was four- to eightfold greater than when the vector was linearized as a replacement vector. By contrast, the ratio of targeted to random integrations at the fah locus did not vary with the linearization site of the vector. The different relationships between the targeting frequency and the vector configuration at the fgr and fah loci may indicate a DNA sequence or chromatin structure preference for different targeting pathways.

UR - http://www.scopus.com/inward/record.url?scp=0027985408&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0027985408&partnerID=8YFLogxK

M3 - Article

C2 - 7969173

AN - SCOPUS:0027985408

VL - 14

SP - 8385

EP - 8390

JO - Molecular and Cellular Biology

JF - Molecular and Cellular Biology

SN - 0270-7306

IS - 12

ER -