Positive-negative selection gene targeting with the diphtheria toxin A-chain gene in mouse embryonic stem cells.
The diphtheria toxin A-chain gene was used in a positive-negative selection gene targeting vector to alter the CD4 gene which is transcriptionally silent in mouse embryonic stem cells. Expression of the toxin gene was driven by a constitutively active enhancer, yet the targeting construct exhibited only minimal transient toxicity while enriching for targeted clones 9- to 29-fold. Germline transmission of the stem cell-derived genome was obtained. These data suggest the usefulness of this diphtheria toxin A-chain cassette in replacement-type positive-negative selection vectors. Its potential for novel applications, particularly in the enrichment for 'hit-and-run' insertion-type vectors, is discussed.
McCarrick
J W
JW
Wistar Institute of Anatomy and Biology, Philadelphia, PA 19104.
Parnes
J R
JR
Seong
R H
RH
Solter
D
D
Knowles
B B
BB
D000818:Animals; D015704:CD4 Antigens / Q000235:genetics; D002460:Cell Line; D004247:DNA; D004167:Diphtheria Toxin / Q000235:genetics*; D004622:Embryo, Mammalian / Q000166:cytology; D005822:Genetic Vectors; D051379:Mice; D016296:Mutagenesis; D010446:Peptide Fragments / Q000235:genetics*; D013234:Stem Cells
D015704:CD4 Antigens; D004167:Diphtheria Toxin; D010446:Peptide Fragments; C029795:diphtheria toxin fragment A; D004247:DNA