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產(chǎn)品名稱：PlatformReagent
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產(chǎn)品廠商：ChromoTek
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PlatformReagent

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Product details
Product details
Purpose	For fast and convenient characterization of interactions between GFP- and RFP-tagged proteins in live mammalian cells by conventional fluorescence microscopy.
Brand	F2H?
Characteristics	transfection supplement enabling assembly of the nuclear PPI-platform

Application Details
Application Details
Comment	Intracellular analysis of protein-protein interactions (PPIs) is crucial for understanding of intimate relationships of proteins within their native cellular environment. Besides, cell-based analysis is often required to support biochemical data obtained with in vitro PPI assays. With the Fluorescent Two-Hybrid (F2H?) Kit, evaluation of selected PPIs can be rapidly carried out in live mammalian cells. Interactions are visualized by simple fluorescence microscopy as co-localization of green- and red fluorescent signals at a PPI-platform in the nucleus of F2H?-Cells, transiently co-transfected with interacting GFP- and RFP-tagged proteins.
Restrictions	For Research Use only

Handling
Handling
Concentration	1 mg/mL
Storage	4 °C/-20 °C
Storage Comment	Store Platform Reagent at +4°C (short term) or in -20°C (long term).
Expiry Date	6 months

Images
Images
Supplier Images	F2H? principle: - F2H cells (BHK origen) carry a GFP-anchoring platform in the nucleu... F2H? result: Exemplary F2H images obtained by fluorescence microscopy Left, inte... An overview image of co-transfected F2H?-Cells at 20X magnification. Cells were PFA-f... F2H?-Cells were co-transfected with interacting proteins, fixed, stained with a nucle... Images for product: Platform Reagent F2H? principle: - F2H cells (BHK origen) carry a GFP-anchoring platform in the nucleus - GFP- and RFP-fusion proteins (bait and prey) are transiently expressed in F2H cells - GFP-bait is recruited to the platform - bright green spot - If RFP-prey interacts with the bait - red fluorescent spot co-localizes with the green spot - If RFP-prey does not interact with the bait - no red spot, disperse red fluorescence F2H? result: Exemplary F2H images obtained by fluorescence microscopy Left, interaction : GFP-bait is recruited to the platform an is visible as a bright green spot. RFP-prey interacts with the GFP-bait and forms a bright red spot co-localizing with the green spot. Right, no interaction: GFP-bait is recruited to the platform and is visible as a bright green spot. RFP-prey is not interacting with the GFP-bait, red signal is diffuse and not enriched at the green spot. An overview image of co-transfected F2H?-Cells at 20X magnification. Cells were PFA-fixed, nuclei were stained with DAPI. Co-transfection efficiency is ~20% in this experiment. Scale bar, 40 μm. F2H?-Cells were co-transfected with interacting proteins, fixed, stained with a nuclear dye (DAPI) and imaged using a 63X objective. Yellow arrows point at the interactions, which are clearly visible at this magnification. Scale bar, 10 μm.

References
References
Product cited in:	Yurlova, Derks, Buchfellner, Hickson, Janssen, Morrison, Stansfield, Brown, Ghadessy, Lane, Rothbauer, Zolghadr, Krausz: "The fluorescent two-hybrid assay to screen for protein-protein interaction inhibitors in live cells: targeting the interaction of p53 with Mdm2 and Mdm4." in: Journal of biomolecular screening, Vol. 19, Issue 4, pp. 516-25, 2014 (PubMed). Wolf, Mantri, Heim, Müller, Fichter, Mackeen, Schermelleh, Dadie, Leonhardt, Vénien-Bryan, Kessler, Schofield, B?ttger: "The polyserine domain of the lysyl-5 hydroxylase Jmjd6 mediates subnuclear localization." in: The Biochemical journal, Vol. 453, Issue 3, pp. 357-70, 2013 (PubMed). Brown, Quah, Jong, Goh, Chiam, Khoo, Choong, Lee, Yurlova, Zolghadr, Joseph, Verma, Lane: "Stapled peptides with improved potency and specificity that activate p53." in: ACS chemical biology, Vol. 8, Issue 3, pp. 506-12, 2013 (PubMed). Wei, Joseph, Sim, Yurlova, Zolghadr, Lane, Verma, Ghadessy: "In vitro selection of mutant HDM2 resistant to Nutlin inhibition." in: PLoS ONE, Vol. 8, Issue 4, pp. e62564, 2013 (PubMed). Mortusewicz, Fouquerel, Amé, Leonhardt, Schreiber: "PARG is recruited to DNA damage sites through poly(ADP-ribose)- and PCNA-dependent mechanisms." in: Nucleic acids research, Vol. 39, Issue 12, pp. 5045-56, 2011 (PubMed).