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Generation of tetracycline-inducible conditional gene knockout cells in a human Nalm-6 cell line

Paper ID Volume ID Publish Year Pages File Format Full-Text
24143 43501 2009 7 PDF Available
Title
Generation of tetracycline-inducible conditional gene knockout cells in a human Nalm-6 cell line
Abstract

Conditional gene knockout by homologous recombination combined with an inducible gene expression system is a powerful approach for studying gene function, although homologous recombination in human cells occurs infrequently. The tetracycline-regulated gene expression (Tet-Off) system is a convenient method for achieving conditional gene knockout, but it is not always promising in Nalm-6, a rare human cell line highly effective for gene targeting. Here we modified the Tet-Off system and applied it to the Nalm-6 cell line successfully by using an internal ribosome entry site to drive a selectable marker from the same tetracycline-responsive promoter for the transgene. We also inserted the gene for the tetracycline-controlled transactivator under the control of a potent CAG promoter. These modifications enabled us to easily obtain rare clones that express optimal amounts of tetracycline-regulated transgenes. We thereby generated a ‘tetracycline-inducible conditional gene knockout’ for the proliferation-associated SNF2-like gene (PASG) in a Nalm-6 cell line, in which the expression of PASG can be depleted in a tetracycline-dependent manner on a knockout background. This method is applicable to any human genes, making this gene-targeting system using the Nalm-6 cell line a promising tool for analyzing gene function.

Keywords
Gene knockout; Gene targeting; Conditional gene knockout; Tetracycline-inducible gene expression system
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Generation of tetracycline-inducible conditional gene knockout cells in a human Nalm-6 cell line
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 141, Issues 1–2, 20 April 2009, Pages 1–7
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us