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Restrictase free generation of targeting vectors for disruption of complex mouse genes.

Abstract：

:Molecular cloning of targeting vectors (TgVs) is a prerequisite procedure for gene disruption in embryonic stem cells. In cases where target genes display complex features (e.g., gene overlap, alternative exon usage), TgVs must mediate deletions with very high precision to prevent unwanted effects. This is often difficult to achieve by procedures using restriction endonucleases and DNA ligases. Therefore, to prepare TgVs for inactivation of two complex genes of immunological interest: PTPRF and NWC, we employed an alternative method, which involves engineering bacterial artificial chromosomes (BACs) by inducible, plasmid encoded "Red/ET recombinase" expression system. Here, we report rapid and efficient construction of PTPRF and NWC TgVs without using restriction endonucleases.

journal_name

Biochem Biophys Res Commun

journal_title

Biochemical and biophysical research communications

authors

Miazek A,Cebula A,Skwarek M,Cebrat M,Kisielow P

doi

10.1016/j.bbrc.2007.04.131

subject

Has Abstract

pub_date

2007-06-29 00:00:00

pages

483-8

issue

2

eissn

0006-291X

issn

1090-2104

pii

S0006-291X(07)00876-5

journal_volume

358

pub_type

杂志文章

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