Document Detail

An Improved Method for the Study of Apoptosis-Related Genes Using the Tet-On System.
MedLine Citation:
PMID:  21335597     Owner:  NLM     Status:  Publisher    
Inducible gene expression systems are particularly useful for the functional characterization of genes with putative toxic properties. In the course of studying the role of hypoxia-regulated gene expression on cell survival using the tetracycline-inducible (tet-on) system, the author noted that exposure to the inducing ligand doxycycline (dox) inhibited caspase-3 cleavage in control samples. to limit this confounding off-target effect, he devised an in vitro pulse dose, delayed-injury protocol testing both dox and a novel tetracycline analog 9-t-butyl doxycycline (9-TB). although 9-TB induced higher transgene levels compared to matched concentrations of dox, continuous exposure to both drugs inhibited caspase-3 cleavage in hypoxic samples. Conversely, a 6-h pulse dose of 9-TB followed by a 40-h washout period prior to hypoxic challenge activated robust transgene expression and lessened the inhibitory effects on caspase-3 processing. It is anticipated that these protocol modifications will improve the performance of tet-regulated genetic screens, particularly in situations where cell death is used as a primary end point.
Marc W Halterman
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Publication Detail:
Type:  JOURNAL ARTICLE     Date:  2011-2-18
Journal Detail:
Title:  Journal of biomolecular screening : the official journal of the Society for Biomolecular Screening     Volume:  -     ISSN:  1552-454X     ISO Abbreviation:  -     Publication Date:  2011 Feb 
Date Detail:
Created Date:  2011-2-21     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  9612112     Medline TA:  J Biomol Screen     Country:  -    
Other Details:
Languages:  ENG     Pagination:  -     Citation Subset:  -    
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