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Showing: 41 - 60 of 255 results
  1. Hot Plasmids - June 2020 - Barcoded CRISPR Library, Sparse Cell Labeling, Calcineurin Reporter, and DNA Staining Dye Alternative

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    Blog Post
    .... https://doi.org/10.1101/788679 A destabilized GFP reporter for calcineurin activity in yeast Article...reporter consisting of a quickly-degraded destabilized GFP variant fused to a 4X CDRE repeat. In addition to... Ca2+ (bottom panels) equipped with destabilized GFP calcineurin reporter (blue) and stained with PI (...death. Image from Diessl et al., 2020. Find the GFP reporter calcineurin activity here Diessl et al.,... They created plasmids for purification of tTALE-eGFP and tTALE-mCherry. In comparison to intercalating...
  2. Opto-Nanobodies: Using Light to Manipulate Cell Signaling and Protein Purification

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    Blog Post
    ...Lab monitored how AsLOV2 insertion into mCherry or GFP nanobodies affected binding of their target proteins...co-localization with a membrane-localized mCherry or GFP protein. With this system, if the OptoNB can bind...more OptoNBs, one against mCherry and one against GFP. Insertion of AsLOV2 into loop 1 always generated....  AsLOV2 Inertion Site mCherry #1 mCherry #2 GFP Loop 1 dark-induced dark-induced dark-induced ...
  3. Using Phosphoserine to Study Protein Phosphorylation

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    Blog Post
    ...TOPO E17TAG GFP and pGEX-6P-1. The pCRT7 NT TOPO vector contains a mutant pSer-containing GFP that can be...during optimization experiments. This version of GFP will contain a phosphoserine at amino acid 17 instead...instead of a stop codon, which will allow full length GFP to be translated), and second, through a gel shift...protein. Up to 80% p-Ser incorporation into E17TAG GFP has been achieved with the Rinehart system (6); however...bacterial strain – Addgene #68306 pCRT7 NT TOPO E17TAG GFP plasmid - Addgene #68295 SupD plasmid – Addgene #...
  4. Fluorescent Proteins 101: Aptamer Fluorophores

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    Blog Post
    ...nature of GFP fluorescence to create an improved fluorescent aptamer system. Knowing that GFP’s  structure... Experiments Learn more about GFP Controlling Protein Activity with GFP Resources at Addgene.org Find...increase its fluorescence, essentially mimicking GFP.  Though the aptamer, called Spinach, required longer...
  5. Plasmids for Endogenous Gene Tagging in Human Cells

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    Blog Post
    ...to endogenously tag a wide variety of genes with GFP. These plasmids are available through Addgene, and...shown in the example above, we inserted a linker and GFP tag at the end of the last exon of the gene to ensure...Cas9 makes at the target site. The whole segment—a GFP tag and 1kb of homologous DNA flanking both sides...repair template. This results in the insertion of the GFP tag precisely where we need it, in this example, ...
  6. CRISPRainbow and Genome Visualization

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    Blog Post
    ...signal to noise ratio of the system, they modified GFP-dCas9 and the gRNA scaffold to enhance complex assembly...the amount of background fluorescence from unbound GFP-dCas9. With these improvements, labeling efficiency...sequences, which recruit fluorescent proteins BFP, GFP, and RFP fused to the respective hairpin binding ...Fluorescent Proteins Control Protein Activity with GFP Resources on Asddgene.org Find Fluorescent Protein...
  7. Multiplex Genome Editing with CRISPR-Cpf1

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    Blog Post
    ...array plus a GFP tag. Four weeks post-infection, there was strong expression of Cpf1 and GFP in the targeted...4 targets when an array of 4 crRNAs, Cpf1, and a GFP tag were transfected on a single plasmid. This is...~75% of neurons were co-transduced with Cpf1 and GFP.  Of these neurons, ~15% had indels at all 3 targeted...lentiviral transduction One vector expresses Cas9, eGFP, and up to 4 gRNAs Each gRNA requires its own promoter...
  8. Fluorescent Proteins 101: Monitoring Cell Mobility Using Fluorescent Proteins

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    Blog Post
    ... to track where GFP labelled B cells were in mounted slices of these organs. These GFP labelled cells ...enabled by fluorescent proteins The optimization of GFP and its derivatives by the late Roger Tsien changed...stimulus. Using an epifluorescence microscope and GFP fusion proteins controlled by promoters specific ...the immune lineage, scientists could easily track GFP expressing cells. For example, by harvesting lymph... the infiltration of neutrophils expressing LysM-GFP (green) into a draining lymph node after infection...Laser include: Blue-green fluorescent proteins like EGFP Yellow-orange fluorescent proteins like TagRFP, ...
  9. Fluorescent Protein Travel Awards - FLiPs and Fluorescent Protein Biosensors

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    Blog Post
    ...lactose released activates expression of GFP from the lac operon. GFP expression can then be used to determine...(including lactose), and a lac operon controlled GFP. When the enzyme breaks down the HMO of interest,...second sensor is a fusion protein consisting of sfGFP fused to a His-tagged ABC transporter known to bind...
  10. Hot Plasmids and Viral Preps - September 2021

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    Blog Post
    ...coding sequence for a green fluorescent protein (GFP). The success of the A-to-G conversion could be seen...through the success of the change in expression of GFP, for the stop codon should have changed from “TGA...cytometry is used to sort populations by mCherry and GFP expression to check for ABE’s. b) Quantification ...
  11. Viral Production at Addgene

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    Blog Post
    ... later, Cre-dependent GFP expression was detected with direct fluorescence. GFP was not detected in the...viral genomes (vg)/cell, pAAV-CAG-FLEX-rc [Jaws-KGC-GFP-ER2] (Addgene 84445-AAVrg) alone at 1.1E6 vg/mL, ...prep # 55632-AAVrg). pAAV-CAG- FLEX-rc [Jaws-KGC-GFP-ER2] was a gift from Edward Boyden (Addgene viral...
  12. What's New in CRISPR - Winter 2018

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    Blog Post
    ...plasmids Tagging human transcription factors with GFP Kevin White’s lab recently deposited a set of CRISPR...plasmids to tag human transcription factors with GFP. As part of the Encyclopedia of DNA Elements (ENCODE...
  13. 28 Hot Plasmid Technologies from 2015

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    Blog Post
    ...both the GFP and the RFP proteins in the nucleus. During the first round of translation, the GFP proteins...Compartment mCherry-Rab5 BFP-Rab5 GFP-Rab5B Vacuolar & Budding Compartment GFP-Rab4B Mitochondria mito-BFP...GTPase in division) GFP-Mff (outer membrane protein) Late Endosome mCherry-Rab7A GFP-Rab7A     ...protein Knock-off, is made up of three components: a GFP protein that binds to the coding region of a reporter...reporter mRNA via a PP7 coat protein (NLS-PCP-GFP) a RFP protein that binds to the 3’ UTR of the reporter...these dCas9s with one of three fluorescent proteins (GFP, BFP and mCherry), the authors were able to visualize...monoclonal antibodies against them, fused them to GFP, and compared them to popularly used epitopes tags...
  14. Multicolor Animals: Using Fluorescent Proteins to Understand Single Cell Behavior

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    Blog Post
    ... from the default GFP (green) to RFP, CFP, or YFP fluorescence. In Brainbow 2.1, GFP is tagged with a ...and activate fluorescence of the downstream OFP, GFP, or RFP. Cells that express the construct but do ...generates cells that randomly express either nuclear GFP, membrane CFP, cytoplasmic YFP, or cytoplasmic RFP...incorporates the highly photostable fluorophores mOrange2, EGFP, and mKate2 (red) to solve the problematically low...
  15. Our Most Popular Blog Posts from 2019

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    ... PS: they’re free. 3. Fluorescent Proteins 101: GFP Fusion Proteins - Making the Right Connection by ...Joachim Goedhart shares some strategies for generating GFP fusion proteins. From considering protein size and...
  16. CRISPR 101: Mammalian Expression Systems and Delivery Methods

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    Blog Post
    ... create a fusion with a reporter protein such as GFP? All of these outcomes can be most effectively achieved...typically used for gRNA. May contain reporter gene (e.g. GFP) to identify and enrich positive cells or a selection...transfer vectors. May contain reporter gene (e.g. GFP) to identify and enrich positive cells. Likely contains...repair by non-homologous end joining (NHEJ) or HDR. EGFP validation of sgRNAs: Can evaluate component delivery...efficacy by cloning in genome target sequence into EGFP reporter. Target DNA reporter system: Can evaluate...
  17. Fluorescent Tagging of Endogenous Genes with SapTrap

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    ...Since the discovery of GFP over 50 years ago, the growing spectrum of fluorescent proteins (FPs) has ...repair pathway, a new user-defined DNA sequence (like GFP) can be inserted at the DSB. CRISPR has allowed scientists...toolkit offers C- and N-terminal tagging vectors (eg GFP, Flag, YFP, Strep, TEV-V5) with either blasticidin...NanoLuc), fluorescent proteins (TagGFP2, TagBFP, TagRFP, and T2A-TurboGFP-PEST), and small epitope tags ...library containing several types of fluorescent (EGFP, tagRFP, mCherry) and nonfluorescent (Halo, SNAP...
  18. Hot Plasmids - November 2023

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    Blog Post
    ...transcriptional activation events as bands of induced GFP-iPAK4 expressed under promoters like the tet-responsive...fluorescent signal in the ticker tape from cFos::GFP-iPAK4 within an hour. Analyzing the ticker tape images...to red at 12 h, and PMA activation induces cFos::GFP-iPAK4 at 3 or 6 h. Scale bar 5 µm. Images adapted...
Showing: 41 - 60 of 255 results