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  1. Scientists Map the SARS-CoV-2-Human Interaction Network

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    Blog Post
    ...selection. Expression is driven by a fairly strong EF1alpha promoter. We have verified expression of correctly-sized...2 virus-human interaction network in which we expressed 26 of the 29 SARS-CoV-2 viral proteins in human...the viral protease, Nsp5. Our set also includes a GFP construct as a control. For easier portability, all...Western Blot for 25 constructs, and confirmed expression via mass spec for two of the remaining viral ...going on there!).  Find the Krogan Lab SARS-CoV-2 plasmids (Note: The Nsp6 construct is undergoing QC and... has been inundated with requests to share our plasmids with other researchers. We have been excited to...Gordon) have been quite busy shipping and sending plasmids to all who request them -- in locations all over...
  2. Chromoproteins: Colorful Proteins For Molecular Biology Experiments

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    Blog Post
    ...high-copy plasmid-borne chromoproteins lost color overnight in liquid culture, though expressing the chromoproteins...for allelic exchange Figure 3: pTOX plasmids expressing amilCP and tsPurple genes. Image: Lazarus...coli BioBrick plasmids (meffRed, eforRed, asPink, spisPink, scOrange, fwYellow, amilGFP, amajLime, cjBlue...contrast, the widely used green fluorescent protein (GFP) gives a bright green fluorescence when exposed to...at the cells on the plate. For example, if your plasmid contains a gene encoding a blue chromoprotein, ...colonies to determine which cells have taken up the plasmid. Of course, other follow up methods like sequencing...gfasPurple) (Figure 2). Figure 2: Chromoprotein expression from bacteria on agar plate. Image: Liljeruhm...
  3. New Norepinephrine Indicators: nLightG and nLightR

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    Blog Post
    ...Resources on Addgene.org Find Biosensor Plasmids Find Optogenetics Plasmids Visit the Fluorescent Protein Guide...Representative images of HEK293T cells and neurons expressing nLightG or nLightR before/after application of...ΔF/F0 heatmaps. White insets indicate surface expression of the indicators over white dashed lines. Scale...contains circularly permuted green fluorescent protein (GFP), and nLightR uses a red fluorescent protein, mApple...accounted for in your experimental design. Find nLight plasmids here! Next steps While nLightG and nLightR greatly...
  4. Bacterial Toxin-antitoxin Systems as Molecular Biology Tools

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    Blog Post
    ...drive high protein expression in vivo. Expression plasmids are often lost without plasmid addiction systems...cells to retain plasmids - if a cell does not inherit plasmid due to improper plasmid partitioning, it...unstabilized plasmids were retained in less than 10% of cells while plasmids containing the plasmid addiction...ensure that plasmids are passed down through generations (Figure 1). These so called “plasmid addiction ...only survive if they retain the plasmid. Since their discovery, plasmid addiction systems have been used...found that if a mini-F plasmid carried a specific 700 bp segment of DNA, the plasmid was propagated better...selection in one plasmid, they were able to create an efficient cloning tool. The plasmid pKSCTMF contains...
  5. Another Pathway into Cells: iTOP

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    Blog Post
    ...stem cells transduced with iTOP resulting in GFP expression. Image courtesy of Diego D'Astolfo. CRISPR... mutations and gene expression changes elsewhere in the genome. Second, expression is by nature transient...control could enter cultured cells and activate expression of a luciferase reporter. CPP was not required...causes a cell to take up, and continued protein expression would require multiple transduction events. For... to binary systems in which transient protein expression has a long-lasting effect, such as Cre- or Cas9...10.1016/j.cell.2015.03.028. PubMed. Find the Plasmids @Addgene: iTOP-tested proteins and reporters...
  6. The Michael Davidson Collection: One-Stop Shop for Fluorescent Proteins

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    Blog Post
    ...colleagues built a Molecular Expressions collection including some 3,300 plasmids along with image galleries...Fluorescent Proteins 101: GFP Fusion Proteins - Making the Right Connection.  mEmerald plasmids and many more! ... resources to go with them. This collection of plasmids is available in an easily searchable format on...Fluorescent Protein Collection webpage. These plasmids are divided into the following categories: Over...list of cell markers all tagged with mEmerald. Plasmids to monitor protein-protein interactions through...Specialty constructs for biosensing. Over 300 plasmid backbones spanning a rainbow of fluorescent proteins...proposition, explained Michelle Baird, who has created plasmids as a technician in the Davidson lab for the last...
  7. 27 Hot Plasmids from 2016

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    Blog Post
    ...lines expressing it. The Church lab fixed this issue by creating a new dCas9-VPR expression plasmid that...transfer vector for in-vivo expression and plasmids optimized for dCas9-VPR expression in drosophila and yeast...Cas9 nickase, and Cas9 activator expressing plasmids that can co-express 1-8 gRNAs. Ordon et al., 2016 tested...Neurosci 2016. PubMed PMID: 26950006 Plasmid ID Plasmid Name Plasmid Type 74309 pcDNA3.0-TRPV4-p2A-ferritin-p2A-mCherry...highlight a subset of the new plasmids in the repository through our hot plasmids articles. These brief articles...features and applications of a partiular plasmid or set of plasmids. We hope that these articles make it ... you to find and use the plasmids you need. You can find all the hot plasmids from 2016 below. With over...
  8. Tips for Screening with Yeast Two Hybrid Systems

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    Blog Post
    ...transforming both fusion plasmids, or mating two single haploid yeast fusion plasmid transformants into a ...Addgene Browse Our Yeast Expression Vectors Find Bacterial Two Hybrid Plasmids Read our Blog Post About...activation. Vary the expression levels of bait and prey proteins - overexpression can lead to false interactions...During this period, proteins were successfully expressed as fusions that retain their individual activities...in this case a reporter gene (e.g. luciferase or GFP), regardless of whether it interacts with a transcriptional...see Figure 2a) (4). These fusions are then co-expressed in a yeast strain that also contains a reporter...bait”, Figure 2b) cannot activate reporter gene expression on its own, but, if the two POI’s interact, this...
  9. Split-BioID: An Improved Method for Studying Protein-Protein Interactions

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    Blog Post
    ...Addgene.org Find plasmids from the Bethune Lab Find Plasmids from the Roux Lab Find Plasmids for FRET ...labeling, BirA* fusion proteins are expressed in cells via plasmids or by the creation of stable cell lines... enriched relative to controls using Split-BioID-GFP and BioID run on six unrelated proteins. One caveat... et al saw different levels of fusion protein expression and biotinylation depending on which terminus... requires 6-24 hours for tetracycline induced expression of the BirA* fusion protein and saturation of... al allowed 24 hours for tetracycline induced expression and biotinylation of interacting proteins. This...Interested in using Split-BioID with your research? Plasmids are available from Addgene! References Schopp...
  10. An “elegans” Approach to Better CRISPR/Cas9 Editing Efficiency

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    Blog Post
    ...drug resistance (Chen et al., 2013), fluorescence (GFP; Tzur et al., 2013), or rescue of mutant phenotypes...step as simple as heat-shocking to induce the expression of a Cre recombinase contained within the knock-in...homology arms to knock in large epitopes, such as GFP (Paix et al., 2014). A further development of this...efficiencies, obtaining as many as 100 independent GFP knock-ins from a single injected animal.  PAM sequence.... elegans. These modular toolkits build single plasmid targeting vectors that encode the gRNA transcript... C. elegans, visit Addgene’s C. elegans CRISPR plasmids page. Future perspectives It is interesting to...using modified Cas9 proteins to regulate gene expression (CRISPRi/CRISPRa), visualize specific genomic...
  11. Hot Plasmids February 2024

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    Blog Post
    ...of the new plasmids, antibodies, and viral preps in the repository through our Hot Plasmids articles.  ...all applications. Addgene’s plasmid collection includes over 10,000 plasmids with a FLAG tag (!), but did...escape. Combining Cas9-PAGE with lentiviral sgRNA expression yielded editing of 80–90% in human cell lines..., Myc and SV40 Nuclear Localization Signals, and GFP. B) Workflow to assess Cas9-PAGE editing with viral... al. 2023 with permission.   Find Cas-PAGE plasmids here!   Zhang, Z., et al. (2023). Efficient engineering...structured illumination microscopy in HeLa cells expressing COX8a-mStayGold. Scale bars 10 μm (main), 1 μm...inspired and informed this section.  Find mStayGold plasmids at Addgene here!   Ando, R., et al. (2023). StayGold...
  12. Plasmids 101: Repressible Promoters

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    Blog Post
    ...to be expressed. These permit exquisite control of gene expression and tracing of gene expression across... basal expression. Co-expression of repressor QS along with QF leads to intermediate expression from QUAS...the Church lab uses this system to control expression of GFP.)  To turn this inducible system into a repressible...transgene permits directed expression of that gene in cells that also express GAL4. Usage of this system...UAS to examine the expression of reporter genes, or to combine reporter expression from one promoter with...basal state, and co-expression of the inducer QF can increase QUAS driven expression by ~3,300-fold in ... Shen. “Controlling gene expression with the Q repressible binary expression system in Caenorhabditis ...
  13. Tips for Using FRET in Your Experiments

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    ...interest: Plasmid Color Expression Description pPROEX Aqua Cyan Bacterial Expresses Aquamarine with...Cyan Mammalian Expresses mammalian optimized Aquamarine mCerulean N1 Cyan Mammalian Express a gene of interest...Cyan Mammalian Expresses mammalian optimized CyPet pCyPet-His Cyan Bacterial Expresses CyPet with C-terminal... Mammalian Expresses mammalian optimized YPet pYPet-His Yellow Bacterial Expresses YPet with C-terminal...Mammalian Express a gene of interest fused to the C-terminus of SYFP2 Clover Green Mammalian Expresses Clover...Clover (a GFP variant) commonly used with mRuby2 pLSSmOrange-N1 Orange Mammalian Express a gene of interest...used pairs to measure FRET. The table below lists plasmids that can be used to create your choice of fluorescent...
  14. Tips for arabidopsis transformation

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    Blog Post
    ...plant plasmid resources at Addgene  Choosing an expression system Choosing an appropriate expression vector....org Find plant plasmids Visit our plant resource page Deposit arabidopsis plasmids with Addgene ...this method include: Cloning and transforming a plasmid into the bacterium Agrobacterium tumeficans - a...options, but here are a couple of my favorites. Plasmids from the Tsuyoshi Nakagawa lab are binary gateway...terminally tagged fusion proteins with fluorescent (YFP, GFP, CFP, RFP, etc.) or affinity tags (HA, FLAG). There...prefer Golden Gate assembly, there are a series of plasmids from the Dinneny lab that additionally allow for...for visual selection of transformants based on expression of RFP in the seed (Emami, Yee and Dinneny, 2013...
  15. Don’t FRET: Bimolecular Fluorescence Complementation Makes Visualizing Protein-Protein Interactions Easy

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    Blog Post
    ...Fluorescent Proteins 101: GFP Fusion Proteins - Making the Right Connection. Cellular expression Whether BiFC constructs... independent of PPIs. Overexpression can be assessed by comparing the expression levels between BiFC constructs...because it relies on a single BiFC pair. Find BiFC plasmids for your experiment at Addgene! How to Design ... steps: fusion protein construction, cellular expression, and quantification (Kerppola, 2006, 2013). These...constructs are expressed transiently by transfection or stably impacts the success of BiFC experiments...this can also be perturbed by supraphysiological expression levels. BiFC quantification This is where BiFC...cannot be used as negative controls because their expression level, stability, and subcellular localization...
  16. Donations from Addgene to Yield Answers for Rare Disease Researchers

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    Blog Post
    ...David Goldstein’s lab, will use GFP-ATP1A3, FLAG-ATP1A3, and ATP1B3 plasmids to explore the functional differences...studying four of those rare diseases will receive plasmids from Addgene worth a total of $5000.  Lowe Syndrome...involving the primary cilia. They’ll put a variety of plasmids to work to explore the effect of various OCRL1...there. This will help us acquire a lot of those plasmids so that we can try many different things in parallel...ATP1A3. The gene encodes a sodium-potassium pump expressed specifically in neurons of the brain. McSweeney...mutations. More specifically, they’ve proposed to use plasmids with modified nickase Cas9 protein in order to...
  17. Fluorescent Proteins 101: Fluorescent Protein Timers

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    Blog Post
    ...fluorescent protein timers When FP timers are first expressed, they predominantly fluoresce one color but slowly...timers (whether as part of protein fusions or expressed independently) originally fluoresce green or blue...displays predictable kinetics over 14 hours of expression in C. elegans embryos. However, this initial ...ideal FP timers: 1. Ratiometric determination of expression time The ratio of mature to immature fluorescence...fluorescence from the FP timer is dependent on the total expression time but independent of protein concentration...Calibration curves correlating this ratio to the total expression time can therefore be used to determine how long... long a pool of FP timers has been expressed, either in whole cells or as a pool of fusion proteins localized...
  18. Lighting Up Cell Signaling with Photoswitchable Kinases

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    Blog Post
    ...Proteins Control Protein Expression with Cre-lox Control Protein Activity with GFP Resources at Addgene.... kinase inhibitors, psMEK1 and ERK KTR-mRuby2 expressing cells were incubated with an inhibitor of choice...feedback. 3. Studying signaling pathways in vivo Expressing photoswitchable kinases in vivo not only allows...used in other model systems, but they need to be expressed in a tissue or organism that blue and violet light...photoswitchable kinases in my experiments? You can find the plasmids from Zhou et al here. In addition, it could be...Addgene.org Find KTR reporter plasmids Find photoactivatable fluorescent proteins in empty backbones in the...
  19. CRISPR 101: Targeting RNA with Cas13a (C2c2)

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    Blog Post
    ...multiplexing; expressing 5 gRNAs simultaneously produces comparable gene knockdown to single gRNA expression. Although... a superfolder GFP fusion to stabilize the protein in mammalian cells. LwaCas13a-msfGFP can mediate cleavage...tracking, modulating splicing, and regulating expression of specifically targeted RNAs. Finally (and perhaps...at this finding through analyses of global RNA expression, cell stress, and RNA size distribution. When...27256883. PubMed Central PMCID: PMC5127784. Find plasmids from this paper at Addgene. 2. East-Seletsky,...) (2016):270-273. PubMed PMID: 27669025. Find plasmids from this paper at Addgene.  3. Gootenberg, Jonathan...28408723. PubMed Central PMCID: PMC5526198. Find plasmids from this paper at Addgene. 4. East-Seletsky,...
  20. Hot Plasmids: Spring 2025

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    ...Explore our empty and EGFP retroviral plasmid set, our bacterial plasmid set for expressing tagged proteins,...of the new plasmids, antibodies, and viral preps in the repository through our Hot Plasmids articles. ...backbones special: Strong expression: A CMV promoter drives robust gene expression in mammalian cells. Flexible...proteins, and stay tuned for mammalian expression and AAV plasmid sets launching later this year!  Find Addgene's... share our very own set of empty, HA-, and EGFP-expressing lentiviral backbones designed right here at...validated viral packaging and strong expression after inserting EGFP and provide step-by-step protocols ... own Lentiviral backbone plasmids here! Figure 2: Addgene cloned EGFP into pAG Lenti CMV N-HA Puro...
  21. Evolution of Brainbow: Using Cre-lox for Multicolor Labeling of Neurons

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    Blog Post
    ...recombination, RFP is expressed. Cre can mediate one of two deletions to allow YFP or M-CFP expression; these deletions...inversion. The Brainbow-2.1 construct can express one of four colors (n-GFP, RFP, YFP or M-CFP.) The construct...research? Check out Brainbow plasmids available from Addgene! Find Brainbow Plasmids @Addgene: Brainbow 1.0...1.0, 1.1, 2.1 plasmids Brainbow 3.0, 3.1, 3.2 plasmids References: Transgenic strategies for combinatorial...editing may be the hot new way to manipulate gene expression, but other gene manipulation systems remain valuable...ways to spatially and temporally control gene expression, especially in in vivo models, and new Cre-lox... of Cre-lox. Check out our previous blog post, Plasmids 101: Cre-lox, if you need a quick primer on how...
  22. Fluorescent Protein Travel Awards - Protein Variants, a Serotonin Sensor, and an Artificial Leaf Replica System

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    Blog Post
    ... be degraded, leading to reduced eGFP fluorescence. Cells expressing the library are then sorted by fluorescence-activated...CUSPER bioreporter such that fluorescent proteins expression is induced prior to inoculation with IPTG. In...control of promoters that control cell cycle gene expression.  Additional Resources on the Addgene Blog ...fluorescent protein topics page Get the basics on GFP Find microbiology blog posts Resources on Addgene.org...Addgene.org Find fluorescent protein biosensors Browse Plasmids from the Tsien lab Browse the Michael Davidson...variant library is monitored by fusing the variants to eGFP followed by an internal ribosomal entry site and...a variant has wild-type levels of abundance, the eGFP will be stable and fluoresce. But if the variant...
  23. Cloning Mammalian Cells with the Agarose Method

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    ...assays to select both over- and under-expressing clones. If GFP-encoding cDNA is co-transfected at a favorable... Pages Browse Fluorescent Protein Plasmids Find Mammalian Expression Vectors ...and PCR, depending on the level of endogenous expression. We highly recommend the use of an electronic...ampules each of 4 clones with different levels of expression of our gene of interest. Advantages of the agarose...isolate individual cell clones with different expression characteristics from a population of transfected...Time CRISPR User) The Challenges of Cell Culture Plasmids 101: Mammalian Vectors Resources on Addgene.org...the Lindberg Lab we often make cell lines that overexpress genes of interest; more recently we have also...
  24. 10 Great Guest Posts We're Thankful For

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    Blog Post
    ...Favorite Fluorescent Protein Oligomerizes in Cells GFP oligomerization can lead to misleading artefacts ...Nathaniel Roquet from the Lu Lab at MIT developed plasmid based systems that report the order in which specific... action. AAV: A Versatile Viral Tool for Gene Expression in Mammals Adeno-associated viruses (AAVs) are... plasmid...
  25. Better Dyeing Through Chemistry & Small Molecule Fluorophores

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    Blog Post
    ...chemistry! The discovery of green fluorescent protein (GFP) sparked a renaissance in biological imaging. Suddenly...advances lead to an obvious question: In this age of GFP and its ilk, why should cell biologists talk to chemists...have two parts: (1) a genetically encoded “tag” expressed as a fusion with your favorite protein and (2)... Generate Fluorescent Protein Fusions Request Plasmids from the Michael Davidson Collection Find Fluorescent...photons. For example, moving from transient overexpression of protein fusions to gene-edited cells can...
  26. Protein Tagging with CRISPR/Cas9: A Conversation with Mendenhall and Myers

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    Blog Post
    ...of the second plasmid can be done in one day in one assembly reaction. This second plasmid saves you a ...alerts when plasmids become available.  We’ve also had some requests for variations on the plasmids we’ve developed...and Myers for speaking with us! Find the Plasmids: Plasmids for protein tagging with CRISPR Additional... available at Addgene. For more details on the plasmids that are currently available along with detailed...several projects and this is one related to gene expression. One goal of the ENCODE project is to look at...system, what we are doing is adding in a second plasmid to target an epitope tag onto the transcription...second vector is a homology directed repair donor plasmid with homology arms and a selection marker. If things...
  27. Targeting HIV-1 with CRISPR: Shock and Kill or Cut it Out?

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    ...activated HIV-1, measured via HIV-1 promoter-driven GFP expression. In cell lines that can produce HIV-1 toxic... gRNAs to target the HIV-1 5’ and 3’ LTRs and expressed them along with Cas9 in the T-cell line 2D10. ...of the LTRs joined together. Clonal Cas9/gRNA-expressing cell populations were also immune to HIV-1 reinfection...cycle progression, or apoptosis with Cas9/gRNA expression. A pooled analysis found no evidence of Cas9 .... For two HIV-1 strains, Cas9/gRNA lentiviral expression significantly reduced the HIV-1 copy number, ...preventing re-infection would require sustained expression of Cas9/gRNAs, which could result in higher rates...the cells infected with HIV-1 only after stable expression of Cas9/gRNA. Viral production is higher in this...
  28. Delivery Methods for Generating iPSCs

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    ... as a donor plasmid containing the transposon, along with a helper plasmid that expresses the transposase...lingering expression of reprogramming factors. Non-replicating Plasmids: Generating iPSCs with plasmid-based...plasmid-based expression requires serial transfection of 1 or 2 plasmids that express the reprogramming factors...reprogramming. Additionally, lingering expression or re-expression of viral transgenes in iPSC-derived cells... generating iPSCs by plasmid transfection where 2 of 11 clones tested had plasmid integration. Another...the dose of plasmid the cells receive over the whole reprogramming period and the plasmid will get diluted...larger plasmids have lower rates of transfection. Minicircles: Minicircles are like mini-plasmids. They...
  29. The Fluorescent Vegetables in Aptamer Soup

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    ...Browse fluorescent protein plasmids and resources See all Jaffrey lab plasmids ... cellular processes by transforming cells with plasmids encoding aptamer-tagged RNA and later introducing...aptamer downstream of a known promoter. Addgene’s Plasmids 101: Aptamer Fluorophores describes the enrichment...aptamers able to bind to a small molecule derivative of GFP called 3,5-difluoro-4-hydroxybenzylidene imidazolinone...leading to conformational changes that control the expression of coding RNA. Spinach-based riboswitches can...SELEX, the RNA pool was cloned into bacterial expression vectors to create a library. Bacteria were transformed...quantify RNA transcription by RNA Polymerase III. By expressing Corn under promoters for transcripts uniquely...
  30. Which Fluorescence Microscopy Techniques is Best for Me?

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    ...red light instead of blue to excite a molecule of GFP. Because red light has approximately half the energy...light, two photons of red light are needed to excite GFP compared to only one of blue. Only at the focal point...infinitely small point of light – think of a single GFP molecule – will appear as a fuzzy 200-300 nm spot...Resources Online Microscopy Primer on the Molecular Expressions Website Other major microscopy manufacturers ... Find Empty Fluorescent Protein Backbones Find Plasmids for Optogenetics ...
  31. Technologies Enabled by NanoLuc® Luciferase

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    Blog Post
    ...Turquoise (CeNL; Plasmid #85199), Yellow (YeNL; Plasmid #85201), Orange (OeNL; plasmid #85202) and Red ... Researchers have been sharing plasmids ever since there were plasmids to share. Back when I was in the...specific strategies and tactics for getting the plasmids he wanted. Addgene makes it so much easier to ...kDa. An example is the CRISPaint-NLuc construct (Plasmid #67178) for use in the system outlined in Schmid-Burgk...reporter was used to sort successfully infected/expressing cells by flow cytometry. The isolated cells were...fluorescent protein that could be co-imaged with GFP using the same wavelength for excitation. The result...CyOFP1-NLuc-CyOFP1; Antares). Direct injection of plasmids for Antares or firefly luciferase into mouse livers...
  32. 15 Hot Plasmids from 2017

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    Blog Post
    ...highlight a subset of the new plasmids in the repository through our hot plasmids articles. These brief articles...articles provide a synopsis of a plasmid or group of plasmids' functions and applications. We hope that... you to find and use the plasmids you need. You can find all the hot plasmids from 2017 below. With over...over 50,000 plasmids, we can't write posts for every great plasmid that comes into the repository, but...write about your plasmids in a future blog post. No time to read? Listen to our hot plasmids segment on the... that restrict expression to this cell type. To this end, they constructed AAV plasmids bearing one of...bacterial and mammalian mCyRFP1 or CyRFP1 expression plasmids, FRET constructs, and CaMKII and RhoA sensors...
  33. Antibodies 101: Immunoprecipitation

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    Blog Post
    ...the IP in parallel with both a sample known to express the protein of interest, such as a sample transiently...include knockout cell lines or tissues that do not express the protein. With this control set, you should ...In cases such as this, an epitope such as c-Myc, GFP or V5 can be used to tag the protein of interest ...interactions of interest. Tagged proteins are typically expressed at much higher levels than endogenous proteins...antibody posts  Resources on Addgene.org: Find plasmids encoding recombinant antibodies at Addgene ...
  34. Hot Biosensors 2022: Year-End Roundup

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    Blog Post
    ...summer or just a bright new plasmid. One area with lots of bright new plasmids is our Fluorescent Biosensors...are generated by inserting a circularly-permuted GFP into an intracellular loop of various G-protein-coupled...Figure 2: iGluSnFR3 displays enhanced brightness, expression, and sensitivity compared with earlier SF-iGluSnFR...SF-iGluSnFR. Image from Aggarwal 2022.   Find iGluSnFR3 plasmids here! Find viral prep here: pAAV.hSyn.FLEX.iGluSnFR3.... 2022, under CC-BY-NC license.   Find JEDI-2P plasmids here! Find viral preps here: pAAV-EF1a-DIO-JEDI.... By fusing ITAM to an RTK of interest and co-expressing a cytosolic ZtSH2 reporter, they observe fluorescence...et al. 2022, under CC-BY license.   Find pYtags plasmids here!   Farahani PE, et al. pYtags enable spatiotemporal...
  35. Overcoming the AAV Size Limitation for CRISPR Delivery

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    Blog Post
    ...markers were not increased compared to a control AAV-GFP. Not only did the AAV-SaCas9-sgRNA constructs mediate...capabilities of full-length SpCas9. This set of plasmids includes AAV-Cas9C-VPR for targeted gene activation...activation. Split inteins have also been used to express base editors. References Chew WL, Tabebordbar ...HBB and CCR5 genes in human cells using compact expression cassettes. Sci Rep 5: . https://doi.org/10.1038...bacterial immune system cleaves bacteriophage and plasmid DNA. Nature 468:67–71 . https://doi.org/10.1038...Addgene.org Check out our CRISPR Guide Find CRISPR Plasmids ...
  36. Antibody Validation for Flow Cytometry

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    Blog Post
    ...with mixed expression, with the expression measurable through an epitope tag such as FLAG/His/GFP. Fluorescent...epitopes. Detection of overexpressed (tagged or untagged) protein When expression plasmids are available, or...flow cytometry, and mRNA expression.   If your protein of interest is expressed at similar levels in ... involves the transient transfection of expression plasmids in a given cell line. For flow cytometry, ...differences between endogenous expression and transfected overexpression, this approach cannot confirm ...caused by overexpression. Additionally, if the cell line used has endogenous levels of expression, this can...preferred. The overexpression approach is therefore best suited to cell lines without any expression of the target...
  37. Brain Initiative Collection

    Type
    Collection
    ...IRES EGFP Chemogenetics expression plasmid Scott Sternson 123308-AAV9 pAAV-hSyn-GRAB_NE1m Expresses the...83895-AAV1 pAAV-hDlx-Flex-GFP-Fishell_6 Cre recombinase-dependent GFP expression in forebrain GABA-ergic...83895-AAV2 pAAV-hDlx-Flex-GFP-Fishell_6 Cre recombinase-dependent GFP expression in forebrain GABA-ergic...83895-AAV8 pAAV-hDlx-Flex-GFP-Fishell_6 Cre recombinase-dependent GFP expression in forebrain GABA-ergic...83895-AAV9 pAAV-hDlx-Flex-GFP-Fishell_6 Cre recombinase-dependent GFP expression in forebrain GABA-ergic...83895-AAVrg pAAV-hDlx-Flex-GFP-Fishell_6 Cre recombinase-dependent GFP expression in forebrain GABA-ergic... Gordon Fishell 83900-AAV1 pAAV-mDlx-GFP-Fishell-1 GFP expression in forebrain GABA-ergic interneurons...
  38. Cre-lox system

    Type
    Collection
    ...48201 CAG-GFP-IRES-CRE Cre and GFP coexpression CAG Retroviral Gage 49054 CAG-GFP/cre Cre-GFP fusion CAG...116879 CAG-Cremyc-2A-GFP GFP and Cre CAG Mammalian Lu 117148 Hiv7CMV-Cremyc-2A-GFP GFP and Cre CMV Lentiviral...dependent DsRed expression Mammalian Cepko 13770 pCALNL-GFP Cre dependent GFP expression Mammalian Cepko...pAAV-FLEX-GFP Cre dependent GFP expression AAV Boyden 37402 Cre Stoplight 2.4 Zsgreen is expressed in cells... Plasmid Collections Cre-lox Plasmids Cre-lox Plasmids Additional...preparations of plasmids that are currently in our repository. ID Plasmid Description Promoter Expression System...iRFP670-EFS:Cre-2A-GFP iRFP670, Cre, and GFP TRE Mammalian Jacks 68544 AAV-Cre-GFP Cre CMV AAV Nestler...
  39. Retrovirus Plasmids

    Type
    Collection
    ...Weinberg 10676 pMKO.1 GFP MoMLV U6-driven plasmid for shRNA expression; also expresses GFP. See pMKO.1 puro...also expresses luciferase Lowe 9044 pMIG MSCV Plasmid for transgene expression; also expresses GFP. Also ...MSCV-IRES-GFP MSCV For transgene expression and GFP marker Reya 21654 pMSCV PIG (Puro IRES GFP empty plasmid...fused to eGFP expression Clevers 18760 MSCV IRES Luciferase MSCV Plasmid for transgene expression; also ... and Packaging Plasmids Common γ-retroviral envelope and packaging plasmids. ID Plasmid Type Description...with packaging plasmid Weinberg 12259 pMD2.G Envelope VSV-G-expressing envelope plasmid Trono 1733 pCI-VSVG...Ecotropic MLV expressing envelope plasmid Nienhuis and Salmon 35616 pCAG-VSVG Envelope VSV-G-expressing envelope...
  40. Adenovirus Plasmids

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    Collection
    ... Viral Plasmids Adenovirus Plasmids Adenovirus Plasmids Additional Resources: Adenovirus...yeast. Two plasmids, termed a shuttle plasmid and an adenoviral (also called backbone) plasmid, are recombined...contains AdEasy-1™ plasmid Vogelstein Adenoviral Plasmids Common shuttle and adenoviral plasmids. Want an adenoviral... in E3 and insertion of GFP expression cassette. Bunz 179202 pAd5-B6/7 ∆E3-GFP Adenoviral A block for ...viruses with multicistronic expression + Venus tag; See plasmid 62622 for plasmid without Venus Wang Adenoviral...Adenovirus Guide Virus Biosafety Lentiviral Plasmids AAV Plasmids Viral Vectors 101 eBook Adenoviruses are...Strain for recombination between shuttle plasmid and adenoviral plasmid Vogelstein 16399 AdEasier-1 BJ5183 ...
  41. Empty Backbones - Choosing Your Perfect Plasmid Backbone

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    Collection
    ...transient expression vector GFP Varies pMKO.1 GFP - Retroviral shRNA expression pLenti CMV...insect expression Fluorescent proteins (GFP, mCherry, etc) Localization pcDNA3-EGFP - C-terminal GFP for ...Species-Specific Expression If you want to drive expression of your favorite gene, you will need a plasmid with...EF1a, CAG, Ubc Transient expression - Gradia Lab plasmids for mammalian expression, Ligation Independent ...TEV cleavage Transient expression - Gradia Lab plasmids for bacterial expression, Ligation Independent ... Expression - Rubin lab plasmids for Drosophila expression pMT-puro - Inducible...for mammalian expression pLV-mCherry - 3rd gen lentiviral plasmid expressing mCherry pBad-EBFP2 - Bacterial...
  42. Brain Armamentarium

    Type
    Collection
    ...Viviana Gradinaru 37825-CAP-B10 pAAV-CAG-GFP AAV expression of GFP from the CAG promoter Edward Boyden Viviana...Viviana Gradinaru 37825-CAP-B22 pAAV-CAG-GFP AAV expression of GFP from the CAG promoter Edward Boyden Viviana... Gradinaru 37825-CAP-MaCPNS1 pAAV-CAG-GFP AAV expression of GFP from the CAG promoter Edward Boyden Viviana... Gradinaru 37825-CAP-MaCPNS2 pAAV-CAG-GFP AAV expression of GFP from the CAG promoter Edward Boyden Viviana... BRAIN Armamentarium Project below. Plasmids ID Plasmid Description PI Viral Preps ID Viral Prep Description...pAAV-AiE0873m_3xC2-minBG-CoChR-EGFP-WPRE3-BGHpA (Alias: CN5140) Expression of CoChR-EGFP in striatal cholinergic...pAAV-AiE0452h_3xC2-minBG-CoChR-EGFP-WPRE3-BGHpA (Alias: CN4035) Expression of CoChR-EGFP in striatal indirect pathway...
  43. Retrograde AAV viral preps

    Type
    Collection
    ... pAAV-mDlx-GFP-Fishell-1 Dlx GFP Control Fishell 83895 pAAV-hDlx-Flex-GFP-Fishell_6 Dlx GFP Control Fishell... PI 37825 AAV-CAG-GFP CAG GFP Control Boyden 51502 AAV pCAG-FLEX-EGFP-WPRE CAG EGFP, Cre-dependent Control...mCherry)-EGFP EF1a Color-flipping switch. Expresses NLS-mCherry in the absence of Cre, and expresses NLS-EGFP...EGFP-tagged Cre expression Recombinases Wilson 105551 pENN.AAV.CamKII.HI.GFP-Cre.WPRE.SV40 CamKII GFP-tagged Cre...Cre expression Recombinases Wilson 105553 pENN.AAV.hSyn.Cre.WPRE.hGH Syn Cre expression Recombinases Wilson...CamKII Cre expression Recombinases Wilson 107738 pAAV-hSyn-Cre-P2A-dTomato Syn Cre expression with simultaneous...simultaneous dTomato expression Recombinases Larsen 107788 AAV.rTH.PI.Cre.SV40 rTH Cre expression Recombinases...
  44. Caltech Systemic Capsids

    Type
    Collection
    ...Dimidschstein 135631 pAAV-S5E2-GFP-fGFP E2 regulatory element GFP, membrane-targeted GFP Control Dimidschstein ...pAAV-CAG-GFP CAG GFP Control Boyden 50465 pAAV-hSyn-EGFP hSyn EGFP Control Roth 50469 pAAV-CaMKIIa-EGFP CamKIIa...104061 CAG-NLS-GFP CAG NLS-GFP Control Gradinaru 105530 pAAV.CMV.PI.EGFP.WPRE.bGH CMV EGFP Control Wilson....hGH GFAP Cre-expression Cre Wilson 107738 pAAV-hSyn-Cre-P2A-dTomato Syn Cre-expression Cre Larsen 140135...Promoter Description Category PI 37825 pAAV-CAG-GFP CAG GFP Control Boyden MaCPNS2 These viral vector preparations...Promoter Description Category PI 37825 pAAV-CAG-GFP CAG GFP Control Boyden CAP-B10 These viral vector preparations...Promoter Description Category PI 37825 pAAV-CAG-GFP CAG GFP Control Boyden CAP-B22 These viral vector preparations...
  45. Fluorescent Protein Guide: Empty Backbones

    Type
    Collection
    ...dimerization pcDNA3-EGFP - Mammalian Expression pCAG-GFP - Mammalian Expression pLV-eGFP - Mammalian Lentiviral...Mammalian Expression mEGFP-C1 - Mammalian Expression mEGFP-pBAD - Bacterial Expression mAzamiGreen 492 505...Mammalian Expression pRSETA-PAGFP - Bacterial Expression tandem PA-GFP-KanR - Yeast Expression pFA6a-PA-GFP-kanMX6...pFA6a-PA-GFP-kanMX6 - Yeast Expression pFA6a-link-yEPAGFP-CaUra3 - Yeast Expression mPA-GFP-N1 - Mammalian Expression...Bacterial Expression iLOV-N1 - Mammalian Expression iLOV-C1 - Mammalian Expression AcGFP1 475 505 28 Prone ...Mammalian Expression mEmerald-C1 - Mammalian Expression mEmerald-pBAD - Bacterial Expression EGFP 488 507...pBad-EBFP2 - Bacterial Expression EBFP2-N1 - Mammalian Expression EBFP2-C1 - Mammalian Expression EBFP2-pBAD - ...
  46. Tetracycline Inducible Expression

    Type
    Collection
    ...of interest tTA expression plasmid Components for a tetracycline on system are: Plasmid containing a tetracycline-dependent... rtTA or TetR expression plasmid Stable cell lines can be made that continuously express a system component...generation lentiviral plasmid for inducible expression of shRNA; neomycin selection; plasmid 21915 has puromycin...gene expression at a single copy in mammalian cells; Expresses firefly luciferase hairpin and GFP under...option rtTA On Kowarz 16623 pBI-GFP Expression of your gene of interest & GFP from a bidirectional tet-responsive...pTet-IRES-EGFP Lentiviral plasmid for inducible expression of transgene of interest and EGFP None Either... and additional plasmids tetR-KRAB On Aebischer & Trono 16542 pBI-MCS-EGFP Expression of your gene of ...
  47. Special Delivery: Fluorophore Targeting for FRET Studies

    Type
    Blog Post
    ...rewards could be substantial: Plasmid tRNA Synthetase Expression Host pMAH-POLY Polyspecific aminoacyl-tRNA...Resources on Addgene.org Find Plasmids for FRET Browse Empty Plasmids to Fluorescent Protein Fusions ...with natural chain termination at stop codons, expression levels of these tagged proteins are often very...oocytes, mainly). Nevertheless, tRNA suppression plasmids are available from depositing labs that have mastered...Fig. 1) which can be carried out using Addgene plasmids. His-tag labeling reagents You may already have...protein, permeabilized cells  Cy5NTAc 32 25-45 GFP (fluorescein is similar)   Lumio Red Tc: CCPGCC 48...
  48. The Challenges of Cell Culture

    Type
    Blog Post
    ...Dangers of Using GFP for Protein Localization Resources on Addgene.org Find Plasmids for Mammalian Cells...along with Down-Regulation of HIF-1α by NDRG2 Expression Attenuates Proliferation and Invasion in Renal...
  49. Bacterial Expression Systems

    Type
    Collection
    ...Controlled Expression Reporter Plasmids Reporter Plasmids Tagging and Visualization Purification Controlled Expression...Reporter Plasmids You may also like… Plasmids 101 Addgene's Molecular Biology Guides Plasmids 101: E. ... on expression only at a specific time. This collection has a variety of plasmids whose expression levels...interest. Plasmid ID Promoter Inducer(s) PI Purpose Murray Lab pBEST plasmids Various Plasmids pBAD, OR2...drive gene expression at a specific level. Return to Top Reporter Plasmids Reporter plasmids can be used...studying gene expression in Gram-negative bacteria. pAK Series Various Plasmids pTac GFP, Luciferase (...Overview of plasmids used for expression in bacterial systems Plasmid...
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