...Chang-Deng Hu Green sfGFP Split super-folder GFP, our most-requested GFP(1-10) and GFP(11) Versatile protein... Red mScarlet Split-wrmScarlet variant of mScarlet Split-wrmScarlet and split-sfGFP: tools for faster...superpositive GFP Split-superpositive GFP reassembly is a fast, efficient, and robust method for detecting protein-protein...Collection Split Luciferase systems, including NanoLuc®, DULIP, LuTHy Other split protein tools: Split-TurboID...cells with split fluorescent protein. Kamiyama et al. Nat Commun. 2016 Bo Huang spGFP Split superpositive... new and popular split-FP plasmids from our catalog to help you find the right split-FP plasmid for your...Capri(1-10) for use with GFP(11) Multiplexed labeling of cellular proteins with split fluorescent protein ...

...self-assembling split GFP (GFP1-9 and GFP10-11) by flipping a single β-strand of GFP, hence the name FlipGFP. FlipGFP...apoptosis reporter, ZipGFP, which utilized the self-assembling split GFP (GFP1-10 and GFP11) to increase fluorescence... How does FlipGFP work? GFP contains 11 β-strands and one central ɑ-helix that can be split into 3 parts...protein-protein interaction sensor based on tripartite split-GFP association." Scientific reports 3 (2013): 2854... superfolder Cherry (sfCherry). Like FlipGFP, sfCherry was split into  β1-9 and β10-11, where β10-11 was...Lab used FlipGFP to track the spatiotemporal dynamics of apoptosis during embryo development. GFP fluorescence...PMC2117903. To, Tsz-Leung, et al. "Rational design of a GFP-based fluorogenic caspase reporter for imaging apoptosis...

...can be found at Addgene. Split FPs – some FPs (e.g. GFP, Venus) can be split into two halves, which are... of GFP, it's variants, and their relevant mutations, check out Marcy's post from last week on GFP. Additionally... a few hours. For example, superfolder GFP (sfGFP) and mNeonGFP can fold in <10min at 37°C, mCherry takes...are PA-GFP, Dendra2, and the mEOS proteins. Some proteins are reversibly switchable (e.g. rsEGFP, Dreiklang...affected by the temperature. For instance, enhanced GFP (EGFP) was optimized for 37°C, and is therefore most... the discovery of the Green Fluorescent Protein (GFP), a protein from the jellyfish Aequorea Victoria....below (or emission wavelenght range).  By mutating GFP, the variants blue FP (BFP), cyan FP (CFP), and yellow...

...to bind GFP. The idea of using GFP as a scaffold suddenly seemed very realistic. Envisioning GFP as a substrate..., CRE-DOG (Cre Dependent On GFP), is activated by GFP and derivatives GFP and YFP, but not red fluorescent...presence of GFP. These systems, known as T-DDOG and Cre-DOG, respectively, repurpose popular GFP reporter...lines expressing GFP in specific cell types to do more than just label cells. If GFP could be co-opted...selectively manipulate only GFP-labeled cells. Once Tang and Cepko found a description of GFP-binding nanobodies...tested pairs of their GFP-binding proteins (GBPs) to find those that could co-occupy GFP. Once they’d found...culture system. In cells cotransfected with GFP and the GBPs, GFP linked the two GBPs together to create a...

...fused to a fast-splicing split intein and co-infected into the cell. Intein splitting removes any exogenous...native peptide bond at the split site. Figure 1: The base editor is split into two plasmids and co-transfected...efficiency, the split editors transduced at levels similar to that of a single AAV expressing GFP. The lab tested...the therapeutic potential for the split base editors. Find the split base editors at Addgene. Levy et ...deposit please sign up here.        AAV delivery of split base editors Article contributed by Jennifer Tsang...overcome this problem, David Liu’s lab developed split cytosine and adenine base editors that can be reconstituted...voltage indicators, however, fall short compared to GFP-based sensors in terms of response and brightness...

...controls using Split-BioID-GFP and BioID run on six unrelated proteins. One caveat of Split-BioID is that...outside the native cell context. The Split-BioID method Enter Split-BioID, a spatiotemporally controllable...within a ~10 nm radius of the bait. In Split-BioID, BirA* is first split into two non-functional fragments....labeled. Figure 1: Using Split-BioID to Identify Protein Protein Interactions. Split-BioID is a protein complementation...localization of the fusion protein. Advantages of Split-BioID Split-BioID’s key advantage over other PPI methods...Important considerations for Split-BioID It’s important to remember that with Split-BioID, and BioID, any protein...version of Split-BioID. BirA* is fragmented at a different site than in Schopp et al. Find these Split-BioID...

...tested. Find FRET plasmids here. Split EGFP: An alternative to FRET, split EGFP has also been used to study...scientists to begin to study GFP and deduce its biochemical properties. Now GFP and its fluorescent derivatives...expressing GFP from cells that are not. Developmental/transgenic uses: Because of its stability, GFP can be...Fluorescent Protein Mutations Relative to Wildtype GFP EGFP F64L; S65T EYFP S65G; V68L; S72A; T203Y mYFP...halves of EGFP undergo folding, maturation, and fluorescence. Biosensors: A wide array of GFP-based fluorescent...from proteins such as Green Fluorescent Protein (GFP) has likely existed in creatures such as jellyfish...derivatives are a staple in the lab. GFP is used in research across a vast array of biological disciplines...

...One way to reconstitute a split protein like Cas9 is to use split inteins. Split inteins are a pair of naturally...challenge that the field has overcome with split AAVs. In general split AAVs break a large transgene in two ...markers were not increased compared to a control AAV-GFP. Not only did the AAV-SaCas9-sgRNA constructs mediate...length, allowing them to be packaged in AAV.   Split AAV approaches  While finding smaller Cas9 orthologs...In 2016, Fine et al developed a proof-of-concept split intein SpCas9 which had modest editing rates in ...full-length SpCas9. In 2016, Chew et al. developed a split intein spCas9-AAV toolbox that retains the gene-...includes AAV-Cas9C-VPR for targeted gene activation. Split inteins have also been used to express base editors...

...detecting protein-protein interactions that uses split fluorescent proteins tethered to potential protein...visualize PPIs. This fluorescent protein (FP) is split into two fragments that individually lack fluorescence...fluorescence microscopy. Figure 1: How BiFC works. A split FP (e.g., yellow fluorescent protein, YFP) is reunited...information about the structures of the tested PIPs. Many split FPs have been validated for BiFC (Kodama and Hu,...check out our article: Fluorescent Proteins 101: GFP Fusion Proteins - Making the Right Connection. Cellular... Hu, 2012). Such controls are necessary because split FPs can associate, albeit at relatively low frequencies...

...considerably faster, with expression levels comparable to GFP during early embryogenesis (Figure 3). mScarlet-I3...through the middle of each embryo are shown, comparing GFP, wrmScarlet (mScarlet), and the new mScarlet3 and... focal adhesion marker paxillin (visualized with GFP). Reprinted with permission from Zhong et al, 2024...shorter than many popular fluorescence markers like GFP (510 nm), allowing co-imaging. And because bioluminescent...light-sensitive systems. Most excitingly, assembly of the split luciferase is reversible, making PILATeS suitable...., Wang, B., Prakash, M., & Dunn, A. R. (2024). Split Luciferase Molecular Tension Sensors for Bioluminescent...

...release. How is this light-mediated? The researchers split the proteolytically active light chain of BoNT/B...high affinity binding to SspB, brings together the split botulinum neurotoxin halves to enable VAMP2 cleavage...can alter motor responses in C. elegans. Find the split BoTN/B plasmids at Addgene. Liu Q et al., Neuron...cells in vivo, the non-immunogenic violet-excited GFP variant Vex. To understand the impact of this gene...

...in this case a reporter gene (e.g. luciferase or GFP), regardless of whether it interacts with a transcriptional...avoid transcriptional reporters altogether; In the split-luciferase system, intein-mediated splicing reconstitutes... this requirement for transmembrane proteins, a split ubiquitin system has been devised (8). References...Kaihara, A., Sato, M., Tachihara, K., Umezawa, Y.l. Split luciferase as an optical probe for detecting protein-protein...

...Figure 5: A) Representative confocal images showing GFP expression from ssAAV5s with hIBA1 or hIBA1a promoters...Quantifications showing high microglia specificity of GFP expression for the indicated ssAAV5s. C) Quantifications...using reconstitution via mRNA trans-splicing. By splitting a large gene into smaller fragments, delivering... (Figure 6).   Figure 6: Illustration of split fluorophore assay to test reconstitution via mRNA...

...developed into a split aptamer that can be used to investigate RNA assembly. The split aptamer is composed...aptamers able to bind to a small molecule derivative of GFP called 3,5-difluoro-4-hydroxybenzylidene imidazolinone...References Alam, Khalid K., et al. "A fluorescent split aptamer for visualizing RNA–RNA assembly in vivo...

...technology CAG-GFP-IRES-CRE (Plasmid #48201) This retroviral vector from the Gage lab encodes GFP and Cre-recombinase...purposes, retroviral vectors have been developed into ‘split-vector systems’ by separating viral genes and transgene...Vectors   Expressing your gene of interest pMKO.1 GFP (Plasmid #10676) Retroviral vector for cloning and...and expressing your gene of interest with GFP expression.  Cloning and expressing RNAi pMKO.1 puro (Plasmid...

... state-of-the-art NIR FPs can be used similar to GFP-like probes by supplying a single gene to cells. ...crosstalk-free three-color imaging with green and red GFP-like FPs (Shcherbakova et al., 2016), as well as ...protein-protein interactions, we recommend using NIR split reporters based on miRFPs (Shcherbakova et al., ...and higher contrast than previously developed NIR split probes (Tchekanda et al., 2014, Filonov et al., ...for miRFPnanos, 35 kDa for miRFPs, and 27 kDa for EGFP) and a possibility to be used as internal tags due... normalization to fluorescence of co-transfected EGFP. e Originally reported as a monomer16, IFP2.0 was...

...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     ... and maybe we can write a blog post about it! Split Cas9 system The Cas9 protein is composed of an n-terminal...bi-lobed architecture of Cas9 to engineer a series of "split" cas9 molecules that cannot function in isolation...form a fully functional Cas9 upon dimerization. Splitting wild-type Cas9 into n-terminal (Cas9(N)-2xNLS)...molecule can cleave the target DNA. The inducible split cas9 approach can also be used for activation of...

...efficiency is not known, this is easily achieved by splitting transfected cells into 3 plates at different densities...select both over- and under-expressing clones. If GFP-encoding cDNA is co-transfected at a favorable ratio...

...dim GFPs The availability of the three dimensional structure of GFP, its brighter mutant S65T-GFP, and...and GFP shRNA. Mice carrying the R26DsRedR; CRUSH-GFP; and Nestin-Cre alleles showed efficient GFP knockdown...functional complementation between two non-fluorescent split GFP fragments. When the two fragments, expressed in...identified a family of GFP proteins in the cephalochordate Branchiostoma floridae, named bfloGFPs. They went on...and demonstrate that they can successfully create GFP fusion proteins with a variety of genes across the...enabled scientists to engineer a wide variety of GFPs with diversity in fluorescence brightness, intensity...Cas9 gene editing. Dr. Masahito Ikawa has created a GFP reporter plasmid for scientists to validate the efficacy...

...Church lab uses this system to control expression of GFP.)  To turn this inducible system into a repressible...complexity of the system, researchers have used split GAL4 architecture or combined the system with Flp...