...Formulated as AAV-DJ for neuronal expression. CAG-FLEX: CAG promoter, FLEXed for expression in Cre-expressing... signaling and regulation. To create iGluSnFR, the Looger lab wedged GFP between two halves of a bacterially...conformational changes lead to an increase in fluorescence intensity from the inserted GFP. After characterizing ...variants come in AAV packaged virus aliquots being deposited at Addgene. Other plasmids and pAAVs can be ordered...hSynap-FLEX: Synapsin-1 promoter, FLEXed for expression in Cre-expressing cells. hSynap-FLP: Synapsin-1 promoter... Biosensors Browse All Fluorescent Proteins Find AAV Tools References Marvin, Jonathan S., et al. “An...generation of iGluSnFR, the Looger lab swapped eGFP for sfGFP to create SF-iGluSnFR. This improved variant has...

...prep pAAV_BiSSTe4_ChR2_mCherry AAV-PHPeB Optogenetics Gordon Fishell New viral prep AAVrg = AAV retrograde... serotypes AAV-hSyn-Ribo-jGCaMP8s AAV1 Biosensor Marianne Fyhn New viral prep AAV-hSyn-Ribo-jGCaMP8m...dTomato_nlsdTomato AAV-PHPeB Control Gordon Fishell New viral prep pAAV_BiCHATe27_ChR2_mCherry AAV-PHPeB Optogenetics...dTomato_nlsdTomato_nlsdTomato AAV-PHPeB Control Gordon Fishell New viral prep pAAV_BiLAMP5e3_ChR2_mCherry AAV-PHPeB Optogenetics..._nlsdTomato AAV-PHPeB Control Gordon Fishell New viral prep pAAV_BiPVe3_ChR2_mCherry AAV-PHPeB Optogenetics...multiple) rAAV-CAG::FLEX-rev:: hM4D-2a-GFP AAV1 Chemogenetics Scott Sternson New viral prep pAAV-CaMKIIa-hM3D...New viral prep pGP-AAV-syn-flex-iGABASnFR2(no bind)-WPRE AAV1, AAV5 Biosensor GENIE Project New viral...

...have created a cassette that will express GeneR only in the presence of Cre. In this example, Cre would...SSRs) Cre and FLP. SSRs bind DNA at target sites to induce site specific recombination events: Cre recombinase...a cell or tissue-specific Cre is provided. Since the FLEx switch ensures Cre-dependent reporter expression...Schnutgen et al. Nat Biotech 2003): In the presence of Cre, GFP expression is lost, allowing for rescued expression...orientation in the presence of Cre. A similar strategy can be employed for "Cre-off" (i.e. your gene of interest...in the sense orientation and Cre is used to flip it to "off"). These Cre-off switches may be referred ...from our Plasmids 101 series, we learned how the Cre-loxP recombination system can be used to induce site-specific...

...vector. AAV hSyn FLEx mGFP-2A-Synaptophysin-mRuby from Liqun Luo’s lab expresses membrane tethered GFP to ...targeted synaptophysin tagged eGFP. AAV phSyn1(S)-FLEX-tdTomato-T2A-SypEGFP-WPRE from Hongkui Zeng's lab...interneurons with AAV-S5E2-GCaMP6f are now available as a ready-to-use viral prep in serotype AAV1. Distinguish...lab are now available as ready-to-use AAV vectors in serotype AAVrg.   ...  After validating this method through monitoring GFP degradation, this protein compartment was used to...cortical interneurons. We now provide ready-to-use AAV vectors expressing GCaMP6f with this promoter. Image...Distinguish dendritic neurites from axonal neurites with AAV vectors expressing cytoplasmic targeted tdTomato ...

...available including: pAAV-hsyn-GRAB_DA2h in AAV9 New GRAB-DA and 5HT sensors New AAV tools to selectively...coding sequence for a green fluorescent protein (GFP). The success of the A-to-G conversion could be seen...through the success of the change in expression of GFP, for the stop codon should have changed from “TGA...cytometry is used to sort populations by mCherry and GFP expression to check for ABE’s. b) Quantification ...from the Dimidschstein Lab More GCaMP8 ready-to-use AAV preps  ...different biochemical secondary energy sources to increase protein yields from 36 μg/mL to 266 μg/mL. The...oxytetracycline enzymes. Next, this system was used to recreate natural biosynthetic pathways. Biosynthetic pathways...

...operator. AAV AAVs are the workhorses of both chemo- and optogenetics. We’re therefore beginning our AAV viral...currently available) and adeno associated virus (AAV, primarily chemogenetics tools for now but with optogenetic...researcher Connie Cepko has been helping us with AAV testing and recently had this to say about the service...preparations to the scientific community. Addgene's AAV preparations have worked well in my lab, including...ll provide a brief rundown of what lentiviral and AAV preps are available below, but you can always head...also have lentivirus for delivering control shRNAs, GFP, and rtTA from the Tet-on system. The first two can... will be available soon. We’re also distributing AAV preps with particular serotypes that can be used ...

...improve their AAV yields. Given the wide range of downstream applications scientists rely on our AAV preparations... of a fluorescent dye and transduction of GFP-expressing AAV particles we did not observe any difference... of hyperosmolarity on AAV production yields had yet to be examined. Increased yields with sucrose supplementation...membrane When packaging adeno-associated viral vectors (AAV) not all serotypes are equal; while some serotypes...sorbitol. As sorbitol is already a common excipient in AAV formulation buffers, this sugar may be a more attractive...alike as an easy and inexpensive method to improve AAV yields. Although we only investigated 3 serotypes... approach will be beneficial for a host of other AAV serotypes; it will be exciting to test this approach...

...capture module (SpyTag-GFP nanobody) assembled on beads. C) Micrograph of H1.8-GFP nucleosomes collected...In MagIC-cryo-EM, target proteins (expressed as a GFP fusion in this proof-of-concept study) are captured...SpyTag-SpyCatcher linkers and spacer proteins ending with a GFP nanobody (Figure 1). This enrichment streamlines ...) 3D model of the obtained structure of the H1.8-GFP-bound nucleosome. Images adapted from Arimura et ...proteins than SpyCas9, allowing them to be packaged in AAV vectors, potentially with multiplexed targets. ...from Neumann et al. 2024. The team showed that AAV-delivered CHARM could silence the expression of prion... (2024). Brainwide silencing of prion protein by AAV-mediated delivery of an engineered compact epigenetic...

...dim GFPs The availability of the three dimensional structure of GFP, its brighter mutant S65T-GFP, and...carrying the R26DsRedR; CRUSH-GFP; and Nestin-Cre alleles showed efficient GFP knockdown in eyes and clonogenic...strain in which Cre expression causes the induction of both DsRed fluorescence and GFP shRNA. Mice carrying...putida and demonstrate that they can successfully create GFP fusion proteins with a variety of genes across...identified a family of GFP proteins in the cephalochordate Branchiostoma floridae, named bfloGFPs. They went on...CRISPR/Cas9 gene editing. Dr. Masahito Ikawa has created a GFP reporter plasmid for scientists to validate ...adeno-associated viral (AAV) system enables greater spatio-temporal control over expression and increases the number...

...injury markers were not increased compared to a control AAV-GFP. Not only did the AAV-SaCas9-sgRNA constructs...Testing AAV-SaCas9 in vivo To test the efficiency of AAV-SaCas9 in vivo, the Zhang lab created an all-...together (~4.2 kb) into an AAV vector is challenging due to its packaging capacity of AAV (~4.7 kb). While this...co-infection of two AAVs. Cas9 orthologs: shorter, but just as potent and specific? The two AAV strategies described...proteins with AAV, at just 1,108 and 986 amino acids, respectively, the size of both is within AAV’s packaging...in length, allowing them to be packaged in AAV.   Split AAV approaches  While finding smaller Cas9 orthologs...that the field has overcome with split AAVs. In general split AAVs break a large transgene in two pieces...

...Introduction to AAV Find AAV Calcium Biosensor Vectors at Addgene ...ones? Many of the available biosensors use a single GFP or CFP/YFP-derived FRET pair. If you need to free... expression, and compatible with constitutive or Cre-dependent expression. These sensors have also been... tissue-specific targeting. By employing various AAV serotypes, researchers can precisely direct the calcium... ready to go. Pro tip: Vectors with ready-to-use AAV preparations are labeled with the green virus particle...3):1057-65. Resources on Addgene.org Addgene’s AAV Guide Fluorescent Proteins: Biosensors Addgene’... aav...

...restraint include an AAV-based sparse labeling system where a limiting amount of AAV are injected into the... the brain of Cre-expressing mice to trigger recombination and expression of a transgene (Lin et al., ...However, this system requires precise titration of the AAV. So far, tools to overcome this challenge in Drosophila...transcription activation systems, such as LexA/p65 or mCD8/GFP, to allow sparse labeling in a variety of fly cell...site upstream of the stop cassette results in decreased efficiency of recombination and preferentially...

... Mammalian shRNA Tools for RNAi! Cre-Lox technology CAG-GFP-IRES-CRE (Plasmid #48201) This retroviral ...retroviral vector from the Gage lab encodes GFP and Cre-recombinase. The Cre-lox system is a technology that can...the pLKO.1 vector can be found here. Cre-Lox technology Puro.Cre empty vector (Plasmid #17408)   This ...lentivirus, Adenovirus, and Adeno-associated virus (AAV). If you are wondering which virus fits you experiments...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...of mouse transgenics. Experimental uses include: Cre-dependent gene expression, gene knockout, and selection...

...manipulations such as injection of precise titrations of AAV or heat-shocking the organism. The SPARC system uses.... https://doi.org/10.1101/788679 A destabilized GFP reporter for calcineurin activity in yeast Article...reporter consisting of a quickly-degraded destabilized GFP variant fused to a 4X CDRE repeat. In addition to... Ca2+ (bottom panels) equipped with destabilized GFP calcineurin reporter (blue) and stained with PI (...death. Image from Diessl et al., 2020. Find the GFP reporter calcineurin activity here Diessl et al.,...fluorescent protein to the DNA. They created plasmids for purification of tTALE-eGFP and tTALE-mCherry. In comparison... Figure 1: Initial steps in the CRISPR-pooled screening using sgRNAiBAR. Image from Zhu et al., 2019.  ...

... ideal for size-limited AAV delivery. PE6c and PE6d are optimized for dual-AAV systems. These variants...transcriptional activation events as bands of induced GFP-iPAK4 expressed under promoters like the tet-responsive...fluorescent signal in the ticker tape from cFos::GFP-iPAK4 within an hour. Analyzing the ticker tape images...to red at 12 h, and PMA activation induces cFos::GFP-iPAK4 at 3 or 6 h. Scale bar 5 µm. Images adapted...enable the creation of CAR-T cells directly within the body. Overall, DIRECTED can increase on-target ...reprogram delivery vehicles like lentiviral vectors, CreVLPs, and eVLPs for in vivo and in vitro applications.... The Ting lab recently made two deposits that increase the utility of this technique.  By engineering...

...adeno-associated viruses (AAVs), will know just how tricky these tools can be to use. AAVs, which can target ... Cumulative AAV preps available in the Addgene repository each year from 2016–2023. B) AAV preps distributed...like viral vectors that deliver GFP.   Figure 3: A) Cumulative AAV preps available at Addgene from...before selecting an AAV can help a researcher make a more informed choice on which AAV to request and/or...feedback on AAVs at Addgene? Suggest a viral vector for our service; leave a comment; or take our AAV survey...the retrograde AAVrg and the systemic capsid PHP.eB.   Figure 4: A) Cumulative AAV preps for the...from the lengthy process of validating and using an AAV in a specific experimental setup. Teams across Addgene...

...action. AAV: A Versatile Viral Tool for Gene Expression in Mammals Adeno-associated viruses (AAVs) are commonly...Favorite Fluorescent Protein Oligomerizes in Cells GFP oligomerization can lead to misleading artefacts ...Cepko’s Lab at Harvard Medical School describes how AAVs function and indicates important things to consider...reminds us that, although fluorescent proteins are incredibly useful, chemical fluorophores can still be used...Dangerous Viruses Contributed by guest blogger Tessa Cressy from the labs of Dr. Elke Mühlberger and Dr. Rachel...CombiGEM and CRISPR for Combinatorial Genetic Screening CombiGEM is a technology that can be used to quickly...barcode small libraries of CRISPR gRNAs to use for screening experiments. Get the full details on how to make...

...protein of interest. The green fluorescent protein (GFP) and mCherry, two of the most common tags currently...the protein of interest. In our studies, we used AAV viral vectors to express hM4Di in neurons of rhesus...years ago (Field et al., 1988). Thus, we injected AAVs carrying hM4Di-HA in the brains of mice and monkeys...article on the Addgene blog: Fluorescent Proteins 101: GFP Fusion Proteins – Making the Right Connection.   ...Roth BL (2007) Evolving the lock to fit the key to create a family of G protein-coupled receptors potently...

... enhanced brain potency of AAV-PHP.eB.    Figure 4: AAV9-X1 and AAV9-X1.1 efficiently transduce... variants AAV.CAP-Mac for systemic gene transfer in primate brains Endothelial-specific AAV delivery Recombinant...an engineered adeno-associated virus (AAV) vector named AAV.CAP-Mac for non-invasive systemic gene delivery...cells in mice lacking LY6A using AAV1-X1 (Chen et al., 2023); then, when AAV-PHP.eB was injected three weeks..., Myc and SV40 Nuclear Localization Signals, and GFP. B) Workflow to assess Cas9-PAGE editing with viral...-01419-x. PMID: 37430038. Endothelial-specific AAV delivery By Cythina Arokiaraj (Gradinaru lab) Another...delivery to and across brain vasculature of systemic AAVs with endothelial-specific tropism in rodents and...

... how Cre-lox recombination works. Advances in Cre-lox recombination have included inducible Cre, such...adeno-associated viral vectors (AAV) available - AAV-EF1a-BbChT and AAV-EF1a-BbTagBY. These constructs ...Cai et al. also created an Autobow construct containing both Cre and XFPs. Cre production drives recombination...continued utility of Cre-lox. Check out our previous blog post, Plasmids 101: Cre-lox, if you need a quick...membrane tethered cyan). Without Cre recombination, RFP is expressed. Cre can mediate one of two deletions...construct, OFP (orange), is expressed when Cre is absent; stochastic Cre excision results in one of three recombinations...system employs Cre-mediated deletion, the design of Brainbow-2.1 cleverly combines Cre-mediated deletion...