...costly to share. What is a RANbody? A RANbody is a fusion between a reporter and a nanobody that’s encoded...J.R., & Yamagata, M. (2018). Reporter-nanobody fusions (RANbodies) as versatile, small, sensitive immunohistochemical...

.... Historically, glutathione-S-transferase (GST) fusion proteins have been immobilized on a reduced glutathione...glutathione matrix. The advantage of a fusion protein is the efficient purification of an already immobilized...for kinetic analysis Proteolytic removal of the fusion protein could provide a more relevant kinetic analysis...produces biotin ligase and a protein of interest fused to maltose binding protein (MBP, New England BioLabs...protein in bacteria and subsequently purify the fusion protein from a bacterial extract. The MBP tag can...using Xa protease. The carboxyl-terminus of the fusion protein includes an AviTag and a 6X HisTag for ...protein we tested, it may be possible with other fusion proteins - but it would remove both the AviTag ...

...Fluorescent Protein Fusions Observing a bright signal for the first time after cloning an FP fusion with your favorite...consequences of misfolded dark FP fusions including erroneous quantitation of fusion protein levels. It remains...misfolding impacts fusion protein function and this could lead to under and overestimation of fusion protein activity...impact of dark, non-fluorescent, and misfolded FP-fusions on quantitative imaging experiments and cell physiology...interest is exciting and rewarding. But, what if an FP fusion with a resident endoplasmic reticulum (ER) protein...troubling, what if a significant fraction of the fusion protein molecules fail to correctly fold? Misfolded...localization and dynamics of fluorescent protein fusions in live cells in real time. Previously, we have...

...engineering of fusion proteins is relatively straightforward. However, generating a fusion while keeping...Creating fusions at the N- and C- termini of your protein of interest The most straightforward fusion protein...use terminology like “C-terminal fusion,” but I find this very confusing because it can mean that the fluorescent...cloning site for generating fusions. For the POI-fluorescent protein fusions, our experience is that a ...For the fluorescent protein-POI fusions, we generate straight fusions, without any linker. When both orientations...example, the APT1-mVenus fusion localizes at the Golgi, whereas the mVenus-APT1 fusion does not (figure 2)....fluorescent protein fusions There are several choices to consider before generating a fusion protein, e.g. ...

... them to a tumor - they used focused ultrasound (FUS) to heat the tumor area, as the image below shows...stable levels for 6 days - after just 15 minutes of FUS.  Similar constructs to the one above can be made...

...events that occur in less that 5% of the population (Fu et al. 2013, Vouillot et al. 2015). Meanwhile, Sanger...sequence requirement at the terminal 3’ PAM site (Fu et al. 2013; Pattanayak et al. 2013). This approach...magazine.scienceforthepeople.org/.     References 1. Fu, Yanfang, et al. "High-frequency off-target mutagenesis...

...guides (19-20bp)?  Small-scale studies (Tsai et al., Fu et al.) suggest 17-19 bp long guides have reduced...PMID: 25513782. Pubmed Central PMCID: PMC4320685. 3. Fu, Yanfang, et al. "Improving CRISPR-Cas nuclease specificity...

...However, Cas9 can be toxic if continuously expressed (Fu et al., 2013). Using IDLVs ensures transient expression...94-007-4899-6_7. PubMed Central PMCID: PMC7120651 Fu, Y., Foden, J. A., Khayter, C., Maeder, M. L., Reyon...

...PMID: 26829887. PubMed Central PMCID: PMC4735761. 6. Fu, Y. et al. High-frequency off-target mutagenesis ...

...10.1016/j.neuron.2010.11.021  Gambarotto D, Zwettler FU, Guennec ML, et al (2019) Imaging cellular ultrastructures...

...him on Twitter.   References Arribere JA, Bell RT, Fu BXH, Artiles KL, Hartman PS, Fire AZ (2014) Efficient...

...://doi.org/10.1016/j.tibs.2023.10.006 Tang, Y., & Fu, Y. (2018). Class 2 CRISPR/Cas: an expanding biotechnology...the different types of CRISPR systems can be as confusing as reading Shakespeare for the first time. The...

...Viral delivery methods include focused ultrasound (FUS), intracorporeal (IC), intracerebroventricular (ICV...widely throughout the organism to target large or diffuse biological structures. Of primary interest here...Stanton, A. C., Lagerborg, K. A., Tellez, L., Krunnfusz, A., King, E. M., Ye, S., Solomon, I. H., Tabebordbar...Stanton, A., King, E. M., Ye, S., Tellez, L., Krunnfusz, A., Tavakoli, S., Widrick, J. J., Messemer, K...

...gene activation when fused to dCas9. In the dCas9-SunTag-VP64 system, dCas9 is fused to a scaffold containing...hook proteins fused to a streptavidin core that are able to retain reporter proteins fused to the streptavidin-binding...interest is fused to an sgRNA used to direct Cas9 to a specific genomic locus. When fused to the sgRNA...fluorescent protein fusions that can be used for monitoring cellular dynamics. These fusion proteins have been...When fused to an RNA of interest, Broccoli-DHFBI fluorescence can be used to visualize the fusion in vitro...or activation, the c-terminal cas9 fragment was fused with FK 506 binding protein 12 (Cas9(C)-FKBP-2xNLS...light-activated CRISPR-Cas9 effector, Polstein and Gersbach fused the light-inducible protein domains CibN and Cry2...

...single fusion protein, as in a FRET biosensor, then the local concentration of FPs around that fusion protein... can lead to polymerization of the fusion protein. A dimeric fusion protein will have two FP binding domains...polymer formation as fusion protein concentration increases. The polymerized fusion proteins that form ... blogger Erik L. Snapp. Stop using EGFP/GFP for fusion proteins! Despite multiple studies in high profile..., there's no problem. You're OK. However, if you fuse your protein of interest (POI) to GFP to study the...noncovalently dimerize. That means that your POI fused to GFP or another fluorescent protein (FP) could...tetramers (i.e. DsRed) [3]. This is a problem for fusion proteins. One of the major applications of FPs ...

...of interest(POI’s) are fused to TAs and DBDs (see Figure 2a) (4). These fusions are then co-expressed ....g. transforming both fusion plasmids, or mating two single haploid yeast fusion plasmid transformants...period, proteins were successfully expressed as fusions that retain their individual activities (1). Researchers...effectively (3). In Figure 1 for instance, the DBD of the fusion protein shown is still able to bind to the upstream...containing the target sequence of the DBD. The DBD-POI fusion (commonly referred to as the “bait”, Figure 2b)... two POI’s interact, this will bring the POI-TA fusion (commonly referred to as the “prey”) in close enough... reconstitutes functional luciferase only when fusions of the two halves of the luciferase protein interact...

... of two fusion proteins: 1) the genomic anchor - an inactive, dead Cas9 protein (dCas9) fused to CIB1;...CIB1 and it was fused to both the N- and C-termini of dCas9; and 2) CRY2FL-VP64, a fusion of full-length...can also be achieved using single-chain fusion proteins by fusing Cas9 to photoreceptors. One prime example...- the CRY2 photolyase homology region (CRY2PHR) fused to a transcriptional activator domain (VP64 or p65...p65). Upon expression in the cell, the dCas9-CIB1 fusion binds to the target DNA sequence as directed by...the guide RNA (gRNA), while the CRY2PHR-activator fusion floats freely, depicted in the figure below. Once...tested a variety of combinations to optimize both fusion proteins. The best performing combination was NLS-dCas9...

...tagged BioID2 for N-terminal fusions and Myc tagged BioID2 for C-terminal fusions. Kim, et al. Mol Biol Cell...molecular biologists to fuse fragments of DNA together in a desired order and express fusion proteins in a target...SpyCatcher fusion that can be anchored to amylose resin for solid-phase synthesis of larger fusions through...to generate mAID fusions to endogenous genes. Cell lines with the appropriate fusions could then be selected...problems by fusing an RBP to the catalytic domain of RNA editing enzyme, ADAR. This RBP-ADARcd fusion protein... to easily create protein expression constructs fused with epitopes or fluorophores, generate expression...CRISPR-Cas9-based tool for targeting CpG methylation by fusing catalytically inactive Cas9 (dCas9) with the catalytic...

...epigenome-engineering tools - zinc finger nucleases and TALENs fused to epigenetic modifiers enabled epigenetic modifications...Researchers showed that TALE-TET1 constructs, which fused a TALEN to the Tet1 demethylase catalytic domain...various promoters. Other researchers additionally fused a TAL effector to LSD1 histone demethylase to demethylate...targeting much easier! With the help of guide RNAs, fusions between Cas proteins and the epigenetic modifier...CRISPR applications direct catalytically dead dCas9 fused to a variety of epigenetic modifiers to specific...Transcriptional activation p300 acetyltransferase dCas9 fused to the catalytic domain of p300 acetyltransferase...has deposited pcDNA3.1-dCas9-MQ1(Q147L)-EGFP, a fusion of dCas9 to a small DNA methyltransferase from ...

...proteins can interfere with fused protein function Under some circumstances, the fused fluorescent proteins ...immunohistochemistry methods, these proteins are routinely fused to ‘tag’ or ‘label’ proteins to facilitate its detection...could affect the properties or trafficking of the fusion protein (Kimple et al., 2013). We found that the...initially used a construct in which the hM4Di was fused to mCherry. To investigate the localization of the...neurons, we next hypothesized that a different tag fused to the hM4Di might improve the transport of this...the influenza virus, introduced to the field of fusion proteins more than thirty years ago (Field et al...also found that this is not generalizable to all fusion proteins. For example, we saw that another type...

... NLuc fused to enhanced green fluorescent protein (GpNluc) and the other consists of NLuc fused to the...produce a series of FP-RLuc8 fusions for multicolor imaging. In all of these fusions coelenterazine intiates...plasmid #85203) NLuc fusions. The Nagai lab has additionally deposited more than 25 fusions of the Enhanced... Many of our technology platforms like HaloTag® Fusion Protein, codon-optimized Firefly luciferase genes...better biosensors. NanoLuc®-fluorescent protein fusions for imaging Fluorescent proteins are fantastic ...the more penetrative orange fluorescent protein fusion (OgNLuc). Both fluorescent proteins can be excited... by Nluc emission and the lab has dubbed these fusions LumiFluor Reporters. The LumiFluor reporter constructs...

... disrupted when GFP is fused to these sequences. When N- and C-terminal fusions are not tolerated the ...heterotrimeric G-alpha subunit fusions that do not tolerate N- and C-terminal fusions (Adjobo-Hermans, 2011)....microscope, and finally (iv) good toleration in fusion proteins. Since GFP is such a well-validated, all-round...interfere with the activity or the localization of the fusion protein. Also, in cases where the protein of interest...interest is relatively small, adding GFP may alter diffusion rates. Smaller genetically encoded probes are .... GFP can only be used for tagging proteins GFP fusion proteins are made by connecting its DNA code to...are defined by a gRNA. Since Cas9 is a protein, a fusion with GFP allows for detection of genomic locations...

...interest fused to the C-terminus of SCFP3A Amber N1 Yellow Mammalian Express a gene of interest fused to ...interest fused to the C-terminus of Amber mVenus N1 Yellow Mammalian Express a gene of interest fused to ...can be used to create your choice of fluorescent fusion protein with your gene of interest: Plasmid ...mCerulean N1 Cyan Mammalian Express a gene of interest fused to the N-terminus of monomeric Cerulean mCerulean...mCerulean C1 Cyan Mammalian Express a gene of interest fused to the C-terminus of monomeric Cerulean mTurquoise2... C1 Yellow Mammalian Express a gene of interest fused to the C-terminus of monomeric Venus pCEP4YPet-MAMM...SYFP2 Yellow Mammalian Express a gene of interest fused to the C-terminus of SYFP2 Clover Green Mammalian...

...A&C-BEmax   Fused cytosine and adenine base editors Grünewald et al., 2020 SPACE   Fused cytosine and... A Cas protein (catalytically dead or nickase) fused to a deaminase that makes the edit A gRNA targeting...the inactive dCas9 (Komor et al., 2016). These fusions convert cytosine to uracil without cutting DNA....converted to thymine through DNA replication or repair. Fusing an inhibitor of uracil DNA glycosylase (UGI) to...editor using a cytidine deaminase from sea lamprey fused to Cas9 nickase (Nishida et al., 2016). Target-AID...minimum efficiency of ~50% and confirmed that the fusion protein retained the PAM properties of the individual...further decrease indel formation 1.5-2 fold, the team fused bacteriophage protein Gam to the N-terminus of BE4...

...orientation of the BirA* fusion matters! Schopp et al saw different levels of fusion protein expression and...BirA, an E. coli biotin ligase. In BioID, BirA* is fused to a bait protein and it biotinylates proteins that...Split-BioID method, each BirA* fragment is then fused to one of two bait proteins that are known to interact...proteins are labeled with biotin. For labeling, BirA* fusion proteins are expressed in cells via plasmids or...C-terminal BiraA* (C-BirA*). These two fragments are fused to two proteins, here proteins Y and Z, that are..., and that result in proper localization of the fusion protein. Advantages of Split-BioID Split-BioID’... remains even if the interaction with the BirA* fused bait proteins ceases. Lastly, Split-BioID and BioID...

... proteins fused to CGBEs Instead of using UNG, the Wei Leong Chew Lab developed two fusions of a CBE with...single-stranded DNA (ssDNA) region, using enzymes fused to Cas9 to alter the exposed ssDNA bases. Thus, ...editors target this “non-target strand.” To avoid confusion, we usually use different terminology when discussing...ABEs, the editor they were using consisted of Cas9 fused to an engineered adenosine deaminase capable of ...polymerase β (PB) (Chen et al., 2021). Both of these fusion constructs improved editing in various cell types...3838/BE_SMART/. Even more new DNA repair proteins fused to CGBEs The David Liu lab also experimented with...smegmatis, termed UdgX, they found three proteins whose fusion improved the CGBE: DNA polymerase D2 (POLD2), exonuclease...

...are described in more detail below: Fusion protein construction Fusing FP fragments to your PIPs requires...determine where to fuse the FP fragments. In the absence of this information, you can fuse each FP fragment...constructing fusion proteins, check out our article: Fluorescent Proteins 101: GFP Fusion Proteins - Making...hard-to-achieve factors. For example, FRET requires that fusion proteins be in close proximity and present in large...fluorescence. When complementary FP fragments are fused to two hypothesized protein interaction partners...more difficult with FRET, as the overexpression of fusion proteins it requires can lead to aberrant subcellular...BiFC experiment generally involves three steps: fusion protein construction, cellular expression, and ...

...Fluorescent protein fusions One of the easiest ways to track a given protein is to fuse it to a fluorescent...Instead of directly fusing a fluorescent protein to your protein of interest, you instead fuse it to the synthetic...any case you would use a traditional FP fusion, a SunTag fusion could be used as well. Pros of SunTag ...system is far from perfect. For example, If your fusion protein is expressed at low levels, you have to...copies of the short epitope GCN4. GCN4 recruits GFP fused to the cognate scFV antibody, which is expressed... 24 kDa for GFP alone. Although traditional FP fusions can also affect protein activity, half-life, or...

...dCas9-KRAB fusion, both targeted to the promoter region. Cas9 activators include Cas9 fused to VP64, VP64...Cas9 for your knockout experiment Generating a no-fuss genetic knockout  Knock-out cells or animals are... editors are dead Cas9 (dCas9) or nCas9 enzymes fused to either cytosine or adenosine deaminases to facilitate...various cargo to a specific DNA locus by directly fusing the cargo to dCas9.  This technique has been exploited...transcriptional repressor domain which is frequently fused to Cas9 for this purpose. dCas9-KRAB plasmids for...simplest of the dCas9 activators consists of dCas9 fused to a single transcriptional activation domain - ... an epitope tagged dCas9 and antibody-activator fusion proteins. The Synergistic Activation Mediator complex...

... same protein of interest tagged with different fusion proteins for different experiments over the years...idea. What if there were a way to just make one fusion protein that switches to any color or tag you want...molecule you choose (Figure 1). Just express the fusion protein and add a specific ligand! A self-labeling...structure is the protein of interest, and black is the fusion tag or labeling system. Image adapted from Lavis...2003, Kai Johnsson’s lab developed a self-labeling fusion tag based on the human DNA repair protein AGT that... Sheetz labs demonstrated in 2005 that proteins fused with E. coli dihydrofolate reductase (eDHFR) could...the tip. 649-mDia1 and 549-CP are both SNAP-tag fusions, purified and labeled separately with different...

...transcriptional activators: some methods rely on fusion proteins while others re-engineer components of...Description CRISPR activation can occur through fusing dCas9 with VP64, a strong transcriptional activation...transcription. This is done through creating a dCas9/VP64 fusion protein engineered with aptamers that bind to MS2...dCas9, SunTag uses a repeating peptide array to fused with multiple copies of VP64. By having multiple...consistently throughout cells. VPR Description VPR fuses a tripartite complex with dCas9 to activate transcription...notable CRISPR activators is that it requires a fusion protein, rather than relying on a two-component... activation. dCas9-CBP In this method, dCas9 is fused to CBP, an histone acetyltransferase domain capable...

...ternary complex formation between the FKBP12F36V-fusion and the ubiquitin proteasome machinery through ...expression of N- or C-terminally FKBP12F36V-tagged fusion chimeras or locus-specific knock-in of FKBP12F36V...running in your lab Confirm the functionality of your fusion chimeras For each protein-of-interest, it is important...of comparing functionality of N- and C-terminal fusions and degradation in a cell line of interest prior... used to evaluate degradation of the FKBP12F36V-fusion chimera of interest. In the cell lines we have ...provide the best confirmation of selective FKBP12F36V fusion chimera degradation. In the cell lines we have ..., Erb et al., Weintraub et al.), assessments of fusion chimera functionality (Nabet et al., Erb et al....

...by splitting Cre and fusing each half with PhyB or PIF6, phytochrome-like fusion proteins from the plant...unmodified enzyme. Each half of the base editor is fused to a fast-splicing split intein and co-infected ...plasmids containing a red-shifted Ace-mScarlet, a fusion of a voltage-sensitive inhibitory rhodopsin (Ace2N...mScarlet emission. Figure 2: Ace-mScarlet is a fusion between the voltage-sensitive inhibitory rhodopsin...soon as red light or PCB is no longer present, the fusion proteins separate, inactivating Cre. This is critical...Cre (CreC) and the N-terminus of Cre (CreN) are fused to the truncated Phytochrome B (PhyBΔ) and the active...

...reversible oligomerization when fused to a protein of interest. Fusion protein oligomerization happens...cryptochrome-interacting basic-helix-loop-helix) protein. After fusing the CRY2 and CIB1 domains to separate proteins ...used to control homo-oligomerization of a protein fused to CRY2, although some studies have shown that this...derivative of the opsin CoChR, termied soCoChR, by fusing a fragment of the kainate receptor subunit to the... the C-terminus of the protein. Unlike the diffuse neuronal expression observed with wild type CoChR, ...which TagRFP-T and Dronpa flanked a PKA substrate fused to the phosphopeptide binding domain FHA. The FHA1...

...repressor) fusion protein, the authors show that the VP16 transcription activation domain fusion increases... in cellular functional imaging and in protein fusions. The Gadella lab has deposited all mScarlet constructs...activity. In addition, proteins can be “caged” by fusion to specific domains thereby rendering them inactive... base editing technique uses dCas9/Cas9 nickase fused to a cytidine deaminase for targeted conversion ...minimum efficiency of ~50% and confirmed that the fusion protein retained the PAM properties of the individual...the B. subtilis libraries Transcription factor fusions elucidate iPSC reprogramming mechanism The Ptashne... efficiency of iPSC reprogramming while the HP1 fusion eliminates reprogramming. These results argue that...

...validate that the fusion protein localizes properly by comparing the signal from the fusion itself to the ...fluorescent tag. Once expressed as a fluorescent protein fusion, it is possible to track your protein of interest...finally degraded (2, 3). Once you've created your fusion protein, it's important to validate its function...always possible, to use inert fluorescent protein fusions to visualize subcellular structures. For example...example, you might use your fluoresent protein fused to a signal sequence that traffics the fluorescent protein...size – it is possible that a large or charged FP fusion can change the subcellular localization of a protein...

...are just one (admittedly large) class of common fusion protein tags. Biochemists and molecular biologists...amassed a considerable molecular tool box of tags and fusion proteins to aid in the expression and purification..., glutathione S-transferase (GST), which can be fused to recombinant proteins for one-step purification...immunoprecipitating, or purifying that protein may be to fuse an affinity tag to it. The FLAG, hemaglutinin antigen...column. Like other affinity tags, a His tag can be fused to either the N- or C-terminus of a protein. Unlike... available, and though adding too many tags and fusion proteins to your protein of interest would eventually...scientists use tags, with a few time-tested tags and fusion proteins as examples. The tables list more common...

...vacuolar/lysosomal and cytoplasmic Rosella. When fused with a mitochondria-targeting sequence, Rosella ...mitochondria, or mitophagy. Rosella can also be fused to a protein of interest to track the autophagy ... This autophagy biosensor contains mRFP and GFP fused to the autophagy-related microtubule-associated ...puncta turning red (GFP negative/RFP positive) upon fusion of an autophagosome with a lysosome. An increase...to more yellow and red puncta, while a block at fusion of autophagosomes with lysosomes leads to an increase...Fluorescent Protein, but following translation, this fusion protein is cleaved to generate one molecule of ...

...Nano-lanterns contain a Renilla luciferase variant fused to an FP; when supplied with a luciferase substrate...applications, including optogenetics, biosensors, and fusion proteins. Solving the problems of low luminescence...Saito et al. described the first Nano-lantern, a fusion of a mutagenized Renilla luciferase (RLuc8) with...Renilla luciferase variant are used to excite the fused fluorescent protein. This technique is very similar...compartments over several minutes using Nano-lantern fusion proteins. To further improve long-term imaging,... as good reporter genes for endogenous protein fusions created using CRISPR/Cas9. eNLs offer high luminescent...

...Monitor dynamics of vesicle fusion and release from the plasma membrane by fusing various vesicle cargoes ...more popular timers (whether as part of protein fusions or expressed independently) originally fluoresce...expressed, either in whole cells or as a pool of fusion proteins localized to a specific region of the ...expressing a single FP-timer protein or protein fusion. This one protein method simplifies cloning and...between models for trafficking to the lysosome by fusing Medium-FT to LAMP-2A, a lysosome-associated membrane...

...turnover rates. Additionally, by engineering FP fusions associated with cellular organelles, scientists...processes, including mitosis, mitochondrial fission/fusion, nuclear import, and neuronal trafficking. Although... pitfalls of overexpressed fluorescent protein fusions is to replace the genomic copy of your gene of ...of interest with a fluorescent protein fusion. CRISPR genome editing can help accomplish this goal. The..., allows users to easily create C-terminal tag fusions of endogenous genes in human cells. To use CRISPaint...users to generate a plasmid containing a direct fusion of the desired gRNA to the U6 promoter, and a second...

... neurons and show that smFPs fusions provided better labels than fusions to 3 or more HA tags. 4. High-resolutions...synapses for instance. Also these tags need to be fused to scaffold proteins in order to be expressed stably...contrast to epitope tags, FPs are bigger and their fusion to a POI can affect protein folding and protein-protein...proteins (smFPs). smFPs are literally composed of FPs fused to multiple epitope tags. Indeed, to create the ...difficult to distinguish using standard antibodies. By fusing N-cadherin to smFPs the Looger team was able to...

... create an abasic (empty) DNA site using either fused or endogenous Uracil DNA N-glycosylase (UNG) enzymes...researchers at HuidaGene Therapeutics guessed that fusing MPG to an ABE would allow it to act on newly-deaminated...nucleobase hypoxanthine + a ribose sugar. Their fusion of wild-type MPG to an ABE was able to edit A →...al., 2023). gGBE consists only of engineered MPG fused to nCas9. First, MPG removes the G base from DNA...invented. CGBE C → G Base Editor Converts C → G by fusing a uracil DNA glycosylase to a CBE; the first type... H. (2023). Programmable A-to-Y base editing by fusing an adenine base editor with an N-methylpurine DNA...

...complexes, scientists can simply express a bait protein fused to BirA* in cells and then incubate those cells ...and allows scientists to more selectively target fusion proteins to specific locations in the cell.   Find...increased with higher expression levels but for many fusion proteins this would lead to protein/organelle aggregation... is an in vivo technique where two proteins are fused to complementary reporter proteins that can only...molecule that can reassemble to restore activity when fused to two interacting proteins (Schopp et al., 2016...Roux, Kyle J., et al. "A promiscuous biotin ligase fusion protein identifies proximal and interacting proteins...

...epitope tag fusions to a protein of interest. The plasmids used to generate these fusions all use the ...of functions via different fusions. Some applications of PUF protein fusions reported in the literature...leader sequence. Using a variety of GFP and RFP fusions as their test genes, the researchers at the BIOFAB...little as 3 days. Abil et al., 2014 show that you can fuse custom PUF proteins to a translational repressor...

...45 kDa tag which can be purified (along with its fusion protein) with amylose resin. The MBP tag, although...Unlike MBP, the GST tag can be cleaved off from the fusion protein while it is still bound to the glutathione...proper folding of the tag, which prevents insoluble fusion proteins from being purified by GST (maybe try ...ions (Nickel, Cobalt, Copper, etc.), allowing His fusion proteins to be purified by single step metal affinity...affinity chromatography. Purification of His fusion proteins from E. coli is generally more straightforward...

...of plasmids each containing an anti-GFP nanobody fused with a different functional element (tyrosine sulfation...described here can be used to target any protein fused with GFP, YFP, mCerulean, or other related fluorophores... He made use of catalytically inactive dCas9 by fusing it to an engineered peroxidase, (APEX) that labels...plasmids, catalytically inactive Cas9 (dSpyCas9) fused to APEX2 is expressed from a Tet/Shield1 inducible...the 20 nm radius surrounding the dSpyCas9-APEX2 fusion.  This Sontheimer lab used this technology, which...

...stable scaffold, part epitope binding region, all fused to a fluorescent protein. “Frankenbody,” like the...variable fragments (scFvs) (Inbar et al., 1972) are fused to mature fluorophores, and can bind and light up...scaffolds specific to different targets, the scientists fused them to sequences called complementarity determining...of interest tagged with the HA epitope, the lab fused a fluorescent protein to the hybrid anti-HA scFvs...sensitive cells such as living neurons, and can be fused to other fluorescent proteins and tags including...

...protein that they are fused to. The researchers constructed 22 different mTFP1 fusions to both the C- and...readout, respectively. mNeonGreen can be used as a fusion tag for a wide range of applications from traditional...mCitrine and mOrange or for live cell imaging of fusions and FRET reporters. mTFP1 is also insensitive to...termini of the fluorescent protein and found that the fusion protein localized to their expected locations. ...

...easier - now, one can generate a fluorescent protein fusion, transcriptional reporter, and loss-of-function...template plasmid to create fluorescent and 3xFLAG-tag fusion proteins. The repair template and Cas9-sgRNA plasmid...loss-of-function allele as well as a promoter-FP fusion for the gene of interest. L1/L2 larvae from these...phenotype. These animals contain a fluorescent protein fusion for the targeted gene. After initial testing, Dickinson...genome. Although they tested fluorescent protein fusions, this system could be used to make many other genome...