...short sequences of up to 100 bp, such as epitope tags, design primers with template binding regions to...combining multiple primers, a C-terminal epitope tag can be moved to the N-terminus of a protein-coding... can be introduced in a gene, while adding a GFP-tag (Figure 3). Achieving multiple modifications requires..., TOP10, and Mach1. No common lab cloning strain has failed so far. What is most important is the transformation... to the 5’ end of the reverse primer. Mutagenesis - Mutagenesis is performed by designing primers astride...modification, from insertions, deletions, and point mutagenesis to complex assembly of multiple genes. Primer...rapid method for recombination and site-specific mutagenesis by placing homologous ends on DNA using polymerase...

...and gram negative bacteria periplasm. Efforts to tag endogenous genes for proteins in these compartments...Munro, Sean, and H. R. Pelham. "Use of peptide tagging to detect proteins expressed from cloned genes:...Avaliable at Addgene Read Our Blog Post on Protein Tags Read Other Posts on Fluorescent Proteins Browse ...enabled numerous cell biology assays. However, there has been a pervasive view that most FPs in a cell will...FP-fusion proteins functionally relative to the untagged protein of interest. To avoid the accumulation...

... from this publication at Addgene. A protein-tagging system for signal amplification in gene expression...nuclease activity, catalytically dead Cas9 (dCas9) has many applications, the most well-known being transcriptional...transcriptional activation/repression. dCas9 has also previously been adapted to fluorescently label a sequence...imaging time for this technique. The NIH Common Fund has made mapping the 4D nucleome a specific priority,...ability to label live cells make this system truly advantageous. This new technique, developed in Thoru Pederson...sensitivity of the technique. One potential solution is SunTag, a synthetic scaffold that can be used to recruit...from Addgene!) Since the signal level provided by SunTag is so high, cells may be imaged under lower illumination...

...randomly into the host genome, it has the potential for insertional mutagensis. The 2nd and 3rd generation ...

...of them containing fluorescent markers or protein tags, and suitable for molecular cloning. Plus, we decided...transformation?  Great news. A subset of our collection has been prepared as part of our new cloning grade DNA...

...function. TULIPs Tunable light-inducible dimerization tags (TULIPs), plasmids allow cloning of protein coding...sequences with GFP-LOVpep, cpPDZ, ePDZb and ePDZb1 as tags. pDawn/pDusk Blue-light photoreceptor for light-...turning on and off the light source. Optogenetics has been broadly applied to study the physiology of the...channel. ChR2 is preferred over ChR1 because ChR2 has higher conductance at physiological pH and trafficks...Depolarizing) ChR Variants. The optogenetic toolbox has been expanded by scientists through both the identification...heterodimerization can reconstitute the protein. This principle has also permitted the design of synthetic two-part transcription...Phytochrome, cryptochrome, and LOV (light oxygen voltage)-based systems have been used in many experimental...

...AAV5  pOTTC1596 - pAAV SYN1 HA-hM3D(Gq) 121538  AAV5  pOTTC1484 - pAAV SYN1 HA-hM4D(Gi) 119741  AAV5 ... vectors for experiments encoding a fluorescent-tagged molecular tool.   Find pAAV-FLEX-tdTomato, pAAV-GFAP104...iGABASnFR GABA sensor (pAAV.hSyn-FLEX.iGABASnFR) has good membrane localization and brightness. The F102G...its F102G variant in AAV1 at Addgene! Archon1, a voltage sensor AAV Historically, electrical activity in...measured with implanted electrodes. The Archon1 voltage reporter allows a way to monitor neuronal electrical...Archon1-KGC-GFP-ER2]) of this membrane-embedding voltage detector. Find Archon1 and its cre-dependent version...pAAV-hSyn-EGFP *Also coming in our new 20 ul trial size! Voltage indicators Plasmid Serotype Name 119036  AAV1...

...pair changes, and the addition of protein tags. It also has the flexibility to modify the E. coli chromosome...non-selectable DNA fragments, such as gene replacements and tags.  The typical frequency of recombinants is 1 positive...recombineering system The lambda red recombineering system has three components (Figure 1): 1) Exo, 2) Beta, and... in the center of the sequence. Since lambda red has a higher recombination frequency when the lagging...activating MMR: 1) use a strain of bacteria that has key MMR proteins knocked out or 2) specially design...recombinant clones If a antibiotic resistance gene has been inserted, recombinants can first be selected...repress expression of exo, beta and gam.  A major advantage of using this method for lambda red expression...

...The Rinehart Lab has developed several tools useful for the incorporation phosphoserine into your gene...68295 GFP E17TAG Plasmid 68296 GFP S2TAG Plasmid 68297 GFP Q157TAG Plasmid 68298 GFP S2TAG/E17TAG Plasmid...Plasmid 68299 GFP E17TAG/Q157TAG Plasmid 53225 MBP-MEK1 S218TAG/S222TAG ( aka. PCRT7 tetR pLtetO MBP-MEK1 XX...68301 MBP-MEK1 S218TAG Plasmid 68302 MBP-MEK1 S222TAG Plasmid 68305 Beta lactamase S68TAG Plasmid 69118...-EFSep) Plasmid 34624 SepOTSα tRNA-Sep ( aka. pCAT112TAG-SepT) Plasmid 68283 SepOTSβ Plasmid 68284 SepOTSγ...69118 E17TAG GFP zeo resistance Strain 34929 BL21ΔserB Strain 52055 EcAR7 Plasmid 112031 pNAS1b split mCherry...

...Plasmids Find Empty Backbones with Fluorescent Protein Tags  ... first publication in 2012, the Nagai laboratory has assembled a collection of multicolored nano-lanterns... low luminescence By itself, Renilla luciferase has a low quantum yield, and when transiently expressed...using chemiluminescent proteins for imaging, but it has precluded widespread adoption of the technology. ...Nano-lanterns and your experiments The Nagai laboratory has a done a wonderful job characterizing Nano-lanterns...described the first Nano-lantern, a fusion of a mutagenized Renilla luciferase (RLuc8) with Venus. This luminescent...

...AAV2 pAAV-CaMKIIa-HA-hM4D(Gi)-IRES-mCitrine 121538  AAV5 pOTTC1484 - pAAV SYN1 HA-hM4D(Gi) 121539 ...for activating and inhibiting neurons. Minimally-tagged DREADD AAV from the Richie lab are especially useful...121539  AAV5 pOTTC1596 - pAAV SYN1 HA-hM3D(Gq) New chemogenetics AAV Plasmid Serotype Name 119741  ... AAV9 pAAV-hSynapsin1-FLEx-axon-GCaMP6s New voltage sensors Plasmid Serotype Name 119036  AAV1...

...fusion protein consisting of sfGFP fused to a His-tagged ABC transporter known to bind to the HMO of interest... dronpa fusions. With these various designs, Xin has been able to develop photoswitchable Intersectin,...Intersectin, Cdc42, Raf1, Mek1, Mek2, Cdk5, and Cas9. Xin has demonstrated the utility of these FLiPs in a variety... of cells that actively produce the HMOs. Fatima has demonstrated the successful production of both types...precisely spatio-temporally controlled as a key advantage over other inducible enzyme technologies. She ...

...surprising discovery that both purified CPP-tagged protein and an untagged control could enter cultured cells ...derived from the same cell type; however, their usage has been limited by technical problems. For instance,...binary systems in which transient protein expression has a long-lasting effect, such as Cre- or Cas9-mediated...effects. Third, protein purification (once a protocol has been established) may be less laborious than viral...used for protein purification must be helping the untagged protein enter cells. Subsequent work found that...stem cells, they found very high recombination percentages (78-79% after two rounds of transduction.) Compared...

...cells according to whether or not they have been tagged with a fluorescent protein or dye. It separates... distinguishing cell populations that have been tagged with different fluorescent proteins or dyes. 3.... cell mobility within these complex environments has never been easy and, for many years, researchers ...Intravital imaging powered by two-photon microscopy has uncovered host-pathogen interactions leading to the...proteins can be used in your experiments, experience has shown that some are more reliable and more efficient...important to finely tune the excitation wavelength as it has been shown that even a small incremental change of...this system called Brainbow, the Ubow mouse strain has been created to fate map Langerhans cells and follicular...

... be selected, since it cleaves at the stop codon TAG. Design your repair template with 55-100 base...mScarlet).   Plasmid ID Plasmid Name Tags 69090 pMJ915 MBP (N terminal); 6xHis (N ...CRISPR genome editing has made it easier to create knockout alleles in a variety of species, including...including creating Cre-dependent conditional alleles, has remained a challenge. Enter Easi-CRISPR: a method...left homology arm guanine (i.e. GG); this design has been shown to increase T7 promoter activity. To avoid...

...into a vector with a large tag at the N-terminus, but you don’t have the tag already included in the vector...you can insert both the gene of interest and the tag sequences into the vector in one step without scars...where the vector that you want to use is very large, has a high GC content, contains a lot of repeats--any...In this blog post, I will go over some advantages, disadvantages, and examples of how scientists are using... common, and for good reason. They offer many advantages over the traditional restriction enzyme cloning...thereof) nor a serious time commitment. Another advantage is that this process makes it easy to generate...will be using. Some of these products may offer advantages that Gibson assembly does not such as increased...

...respectively. mNeonGreen can be used as a fusion tag for a wide range of applications from traditional...single-molecule super resolution imaging. mNeonGreen also has little sequence identity to other common fluorescent...fold brighter than EGFP and is photostable. It also has narrow excitation and emission peaks (max excitation...was then screened through successive rounds of mutagenesis for the brightest teal fluorescent proteins that...scientists from Florida State University, they used a mutagenesis approach and found that specific amino acid changes...

...artifacts, it has better folding and maturation kinetics than other dark fluorescent proteins, it has good signal...encoding alternative promoters and/or fluorescent tags are available at Addgene. Shemesh OA, et. al. ...Technology (KAIST) and Institute for Basic Science (IBS) has developed another useful optogenetic tool that enables...facilitate future work with this goal, the Barrick lab has deposited the pDCAF3-IS5 plasmid containing a synthetic...operon originates from Pseudomonas putida CBB5 and has been tailored for expression in E. coli. This operon... contributed by Christina Mork The Murakoshi lab has developed two new FLIM-FRET acceptors, ShadowG, a... due to its very low quantum efficiency. ShadowY has a lower quantum yield and higher absorption than ...

...used for PL were APEX2 and BioID. APEX2 is fast, tagging proximal proteins within minutes, but requires ...being targeted via RNAi. Recently the Vellai lab has created an improved RNAi vector that leads to significantly...sequence of the toehold switch. For example, Green has also developed toehold switches to detect Zika virus...protein motifs than previously thought. Sidhu’s group has made several hundred SH3 domain containing plasmids.... In addition to these constructs, the Sidhu lab has made hundreds of additional plasmids available through...infect millions annually. Because it is highly contagious, it is essential to contain outbreaks when they...

... expressing fluorescent proteins will be used to tag individual strains of bacteria. Their interactions... S. University of Baroda, India  Devarshi Gajjar has 10 years experience in the field of bacterial and...biomedicine and astrobiology. During the last 11 years he has studied various aspects extreme life on earth including...educational resources for scientists globally, Addgene has partnered with various community organizations and...