...Mammalian Cells Learn about the Dangers of Using GFP for Protein Localization Resources on Addgene.org...line identity and purity making it easier for comparable studies to be performed on a cell line even decades...through working for a variety of pharmaceutical companies including GSK, AstraZeneca-MedImmune, and Infors...

...SYFP2 Clover Green Mammalian Expresses Clover (a GFP variant) commonly used with mRuby2 pLSSmOrange-...to target mTurquoise2 to various subcellular compartments pCEP4CyPet-MAMM Cyan Mammalian Expresses mammalian...Clover pGWF1 Cyan & Yellow Bacterial Gateway-compatible vector to express a gene of interest fused between...

...used this cassette to knock out genes, knock in a GFP cassette, and introduced site-directed mutations,...populations to retain more of these plasmids as compared to those without the restriction modification ...

...Vectors Read about Reporter Gene like Luciferase and GFP Resources on Addgene.org Find Plasmids for Your...promoter present in your plasmid backbone must to be compatible with the type of RNA that needs to be made: if...

...liver injury markers were not increased compared to a control AAV-GFP. Not only did the AAV-SaCas9-sgRNA constructs...editing is the potential for off-target effects. To compare the off-target effects of SpCas9 and SaCas9, Zhang...had modest editing rates in HEK-293T cells when compared to the full-length SpCas9. In 2016, Chew et al...cleavage of HBB and CCR5 genes in human cells using compact expression cassettes. Sci Rep 5: . https://doi....

...aptamers able to bind to a small molecule derivative of GFP called 3,5-difluoro-4-hydroxybenzylidene imidazolinone...identified through microfluidic-assisted in vitro compartmentalisation for their high binding affinity to TO1-biotin...of these variants exhibit increased brightness compared to Mango, and all are photostable and useful for...

..., Hess et al. successfully evolved wild type GFP to EGFP using CRISPR-X and subsequent FACS sorting. They...with unique tissue-specific promoters expressing GFP, tetracycline response elements, and shRNAs many ... of DsRed connected to a pH-sensitive variant of GFP (SEP) by a 9 amino-acid linker (see Figure 1). The...several GMAP-compatible vectors for a wide variety of biological applications. First, GMAP-compatible backbones...including a stop codon) formats. The collection is compatible with both standard Gateway cloning and the FNLCR...sites. For instance, the authors generated a GMAP-compatible backbone containing Rosa26 homology arms. The...GMAP, there will be a continuous expansion of compatible backbones, promoters, and genes available to ...

... includes things like viral vectors that deliver GFP.   Figure 3: A) Cumulative AAV preps available..., and jGCaMP8 viral preps distributed per year compared to other AAVs preps from 2016–2023. B) Inset showing...

...are generated by inserting a circularly-permuted GFP into an intracellular loop of various G-protein-coupled...enhanced brightness, expression, and sensitivity compared with earlier SF-iGluSnFR. Image from Aggarwal ...size and reduced sensitivity to pH and calcium compared to previous lactate biosensors, and its affinity...

...fluorescent protein that could be co-imaged with GFP using the same wavelength for excitation. The result... a good story (1). From a deep sea shrimp to a compact powerhouse of bioluminescence, NLuc is 100-fold...yield, brightness and improved maturation time compared to similar orange fluorescent proteins. In their...more than 2-fold brighter signal from Antares as compared to Fluc in this model. The construct for Antares...transfer of energy from one fluorescent protein to a compatible fluorescent protein. FRET can be measured by ...

...Church lab uses this system to control expression of GFP.)  To turn this inducible system into a repressible... cells and ~24,000-fold in human HeLa cells as compared to basal expression. Co-expression of repressor...

...labeling. Size is also a factor. Bulkier tags, such as GFP, may interfere with protein expression or function...realizing it. Yet, this selection is crucial as it will impact downstream outcomes of an experiment, including...increased versatility in probe attachment, and compatibility with far-red and near-infrared fluorophores....Grimm & Lavis, 2022). These variations directly impact the way you can utilize these dyes. The most-used...may prove challenging to stain intracellular compartments in in vivo model systems, so extra care must...emission of labeled agonists and antagonists, impacting the interpretation of experimental results. It's...dopamine D2/D3 receptors (Allikalt et al., 2020) Impact of labeling process: The process of dye labeling...

...measurable through an epitope tag such as FLAG/His/GFP. Fluorescent protein tags, which correlate closely...intracellular labeling because the intracellular compartment may contain cross-reacting antigens not present...difference in the efficacy of your antibody. When compared, 4% PFA and 0.1% saponin; 4% PFA and 0.1% Triton...intensity in the KO population (pink histogram) compared to the WT cells (green histogram). The unfilled..., like RNAseq or proteomics, it is possible to compare antibody labeling of different cell types within...expression of the target. In this case, you could compare the antibody’s labeling of the cell lines to the...

...cassette comprised of OCT4, SOX2, LIN28, NANOG, and a GFP reporter. Minicircles are removed from cells by dilution...efficiency of reprogramming when compared to other methods, even when compared to integrating viral approaches...heterogeneous iPSC cell lines, which can complicate comparisons made between lines. Incomplete silencing of transgenes...and have a higher efficiency of generating iPSCs compared to other delivery methods. Retrovirus has a cloning...lentivirus has similar rates of reprogramming when compared to MMLV-derived retroviruses, it is not as well...heterogeneity between clones. This can complicate comparisons made between samples, i.e. transgene expression...Non-integrating methods have a smaller genetic footprint compared to integrating approaches. These methods eliminate...

...-positive assay) as well as stabilization (Nanog-GFP reporter activation and silencing of factor-linked... Listen to Our Tol2 Gateway-Compatible Toolbox Podcast Segment The impact of KLF4 N-terminal variants ...lead to comparable experimental results. Using PiggyBac transposons, the Woltjen Lab compared different...along with EGFP while retaining an emission spectrum that is easily separable from that of EGFP. This makes...latest hot plasmid updates!     Tol2 Gateway-compatiable toolbox for studying the nervous system The... Cole lab recently deposited the Tol2 Gateway-Compatible Toolobox, which will allow scientists to quickly... neuorodgenerative diseases. The Tol2 Gateway-Compatible Toolbox is based on the original Tol2kit generated...

...fluorescent protein (EGFP) reconstitution. (a) Scheme of validation for DSB mediated EGFP expression cassette...halves of EGFP to recombine by homology directed repair, and resulting in the expression of EGFP. Using ...ordering the sgRNA is simple and easy. Several companies supply gRNA sequences, such as GenScript, ThermoFisher...order them yourself through whatever DNA supply company you currently use. You can also find many previously...strand annealing) took place and reconstituted the EGFP expression cassette. (b) pCAG-EGxxFP target plasmid...pCAG-EGxxFP target plasmid contains overlapping 5′ and 3′ EGFP fragments under the ubiquitous CAG promoter. The...can be placed in multi-cloning site (MCS) between EGFP fragments. The pX330 plasmid contains humanized ...

...biological properties of new fusions can be compared with mEGFP or mTurquoise2, which are established true...fluorescent protein. In the jellyfish Aequorea victoria, AvGFP forms a homodimer. In corals, the red fluorescent...detect weak dimerization tendency, as is present in EGFP. Constantini et al, (2012) developed a cell-based...fluorescent protein is monomeric without the accompanying data to prove it is worthless. The OSER assay...that you will be using. If you do a head-to-head comparison right, it will be valuable information for the... are engineered can be shared to simplify the comparison by others. Together, we can build a valuable ...

...protein they tested — E. coli expressing good ol’ EGFP — showed a slight but detectable change in fluorescence...better, multiple other labs have reproduced their EGFP findings since the initial report in 2023.)   ...   Figure 1: Fluorescence intensity of EGFP-FlavinTag in E. coli changes in response to a ~25 mT...proteins display a similar effect (many do). Fusing EGFP to the self-labeling FlavinTag made a reliable one-component... the proteins that had shown magnetic responses: EGFP, mScarlet, and AsLOV2. After several rounds of semi-random... semi-random mutagenesis and screening, the EGFP and mScarlet variants were showing no obvious signs of...-up studies. “We don’t have a vision of passive impact,” he says, noting that his team regularly shares...

...enhancer regions (Mendenhall et al., 2013). By comparing gene activation when enhancers were active or ...increased gene activation at the targeted site compared to other Tet1 delivery systems. Transcriptional...) Vlatka Zoldoš’ lab has deposited pdCas9-DNMT3A-EGFP and pdCas9-DNMT3A-PuroR for targeted cytosine methylation...methylation in mammalian cells. Co-expression markers EGFP and PuroR enable sorting and selection of transduced...Goodell's lab has deposited pcDNA3.1-dCas9-MQ1(Q147L)-EGFP, a fusion of dCas9 to a small DNA methyltransferase...epigenome-editing tools. pLV hUbC-dCas9-MQ1(Q147L)-EGFP is also available for lentiviral transduction. Lysine-specific...

..., and disease. In order to expand their model’s impact, organoid users have recently begun to develop ...circularly permuted green fluorescent protein or cpGFP) to the petunia SL enzymatic receptor DAD2. SLs ...which modulates the fluorescence intensity of the cpGFP without altering that of the internal control (Fig...these novel biosensors could have a major economic impact.   Design and mechanism of action of the...