Invitrogen™ GeneSwitch™ Mammalian Expression Kit, complete kit

Description

• Expression vector pGene/V5-His provides minimal promoter, GAL4 -E1b, consisting of binding sites for yeast Gal4 DNA binding protein followed by TATA sequence from Adenovirus E1b promoter Without additional factors, GAL4 -E1b promoter is transcriptionally silent To activate transcription from GAL4 -E1b promoter, GeneSwitch regulatory protein is expressed from minimal TK promoter on pSwitch vector GeneSwitch protein has three functional domains: Gal4 DNA Binding Domain (Gal4-DBD) to bind regulatory protein to GAL4 -E1b promoter; Truncated human Progesterone Receptor Ligand Binding Domain (hPR-LBD) that undergoes a conformational change when it binds the progesterone antagonist, mifepristone; Transcription activation domain from NFØB transcription factor p65 (p65AD) to activate transcription from silent GAL4 -E1b promoter In absence of mifepristone, conformation of hPR-LBD region prevents GeneSwitch regulatory protein from activating transcription from GAL4 -E1b promoter When mifepristone is added and binds hPR-LBD region, GeneSwitch regulatory protein assumes conformation that permits to stimulate transcription from GAL4 -E1b promoter Induction of GeneSwitch System leads to activation of gene of interest on vector pGene/V5-His In addition, four Gal4 binding sites upstream of minimal HSV TK promoter on pSwitch can bind pSwitch regulatory protein Therefore, adding mifepristone up-regulates production of regulatory protein Increased levels of GeneSwitchregulatory protein result in induction of gene of interest from pGene/V5-His to levels that can approach those of viral promoters Provides expression vector pGene/V5-His with minimal, synthetic GAL4 -E1b promoter that is transcriptionally silent until activated In addition, GeneSwitch protein upregulates its own expression, leading to amplified expression of gene of interest from pGene/V5-His C-terminal V5 epitope tag for detection with Anti-V5 Antibodies C-terminal polyhistidine (6xHis) tag for rapid purification with nickel-chelating resin and detection with Anti-His(C-term) Antibodies Zeocin resistance gene for effective selection in both mammalian cells and E
• coli Mammalian Expression, Protein Expression, Proteins, Expression, Isolation and Analysis, Regulated Expression