...swapping and generating N and C terminally tagged fusion proteins with fluorescent (YFP, GFP, CFP, RFP, ...transcriptional and translational dynamics of genes and proteins in living plants, to complementing mutant phenotypes...potato contains Agrobacterium T-DNAs with expressed genes: An example of a naturally transgenic food crop....simplifying plant genetic engineering Using arabidopsis in the classroom Resources on Addgene.org Find plant... transformation. There are many motivations to generate transgenic Arabidopsis, from studying transcriptional... floral drip or dip transformation method. The general steps for this method include: Cloning and transforming...Glazebrook, 2002). Find plant plasmid resources at Addgene  Choosing an expression system Choosing an appropriate...

...negative bacteria periplasm. Efforts to tag endogenous genes for proteins in these compartments using CRISPR ...tagging to detect proteins expressed from cloned genes: deletion mapping functional domains of Drosophila...Lindsey M. Costantini, and Erik L. Snapp. "Superfolder GFP is fluorescent in oxidizing environments when targeted...quantitative imaging experiments and cell physiology in general. Pitfalls of Fluorescent Protein Fusions Observing...cellular compartments other than the cytoplasm. Generally, FPs evolved in or were engineered for use in ...N-glycosylation consensus sequences in popular FPs. This generated a palette of inert green, cyan, yellow, and blue...standard FPs. The moxFPs are now available through Addgene Many thanks to our guest bloggers Erik L. Snapp...

...check out our article: Fluorescent Proteins 101: GFP Fusion Proteins - Making the Right Connection. Cellular...been generated for yeast (Kim et al., 2019) and fruit flies (Bischof et al., 2018), the generation of such...quantities (and the correct stoichiometric ratios) to generate useable data. What’s more, measuring and quantifying...straightforward nature of BiFC also makes it easier to generate a large number of constructs for high-throughput...screening of PPIs. For example, Bischof et al. (2018) generated an open reading frame (ORF) library to screen ... such resources in less genetically tractable organisms lags behind. BiFC can also be used to screen for...pair. Find BiFC plasmids for your experiment at Addgene! How to Design a BiFC Experiment Much like FRET...

...which must be stabilized by monomeric superfolded GFP and averages only ~50% RNA knockdown. In initial ...plasmids from this paper at Addgene. Additional Resources on the Addgene Blog Check out our CRISPR Featured...and designated this system REPAIRv1. Using next-generation sequencing, they confirmed A->I editing and found...cytosine deaminase. Three rounds of rational mutagenesis on the ADAR2DD residues that contact the RNA ...RNA-sequencing, the authors performed rational mutagenesis and identified that a S375A mutation in RESCUE...PMID: 29070703. Find plasmids from this paper at Addgene. Abudayyeh, Omar O., et al. “RNA targeting with...PMID: 28976959. Find plasmids from this paper at Addgene. Abudayyeh, Omar O., et al. "C2c2 is a single-...

...In cases such as this, an epitope such as c-Myc, GFP or V5 can be used to tag the protein of interest ...resources on the Addgene blog: Introduction to Immunofluorescence View all of Addgene's antibody posts  ...Resources on Addgene.org: Find plasmids encoding recombinant antibodies at Addgene  ...that interact with nucleic acids. Labs studying epigenetics routinely use the chromatin IP (ChIP) technique... found this blog helpful be sure to check out Addgene’s blog for other antibody topics.     References...

...about the Dangers of Using GFP for Protein Localization Resources on Addgene.org Find Plasmids for Mammalian...pharmacology (many of which are represented in Addgene's Special Collections). Arguably, oncology has benefited...examining whether the cell lines had isoenzymes and genetic polymorphisms specific to their origins. This technique...out by the commercial repository. Repositories generally ensure cell line identity and purity making it...to the original as possible and so any results generated are reproducible and reliable. When you order ...Abcam Cell Culture Guidelines Resources on the Addgene Blog Learn All about Mammalian Vectors Read up...

...varies across targets, they tested two genes in mice. For both genes, they saw indel formation and phenotypic...in vitro and germline genome editing, but in vivo gene editing approaches have been limited by problems...viral vectors (AAV) are commonly used for in vivo gene delivery due to their low immunogenicity and range...markers were not increased compared to a control AAV-GFP. Not only did the AAV-SaCas9-sgRNA constructs mediate...al. successfully used CjCas9 with AAV to target genes in mouse muscle and eye tissue. Neisseria meningitidis...cell is transduced by both AAVs, the full length gene and/or protein is reconstituted. One way to reconstitute...split intein spCas9-AAV toolbox that retains the gene-targeting capabilities of full-length SpCas9. This...

...vectors that deliver GFP.   Figure 3: A) Cumulative AAV preps available at Addgene from 2016–2023 by...in the Addgene repository each year from 2016–2023. B) AAV preps distributed per year by Addgene from 2016...vector preps contain biosensors, optogenetic tools, and chemogenetic tools: protein tools that are activated...biosensor, by Addgene’s categorization) was first developed in 2001, with successive generations developed...As part of Addgene’s 20th anniversary celebration, we’ve been finding and sharing stories unique to our... our repository and resources. As editor of Addgene’s eBooks — including the recently released Viral Vectors...viruses engineered to be safely used to deliver a genetic payload to cells, and the complexity of effectively...

...aptamers able to bind to a small molecule derivative of GFP called 3,5-difluoro-4-hydroxybenzylidene imidazolinone...If you’re been following Addgene on #souptwitter or if you just enjoy a warm meal on a winter day, you...molecule into the cell. The fluorescence that is generated allows for qualitative and quantitative measurement...placing the aptamer downstream of a known promoter. Addgene’s Plasmids 101: Aptamer Fluorophores describes the... we’ll take a deeper dive into the content of Addgene’s aptamer soup by exploring the development and ...and applications of fluorescent RNA aptamers in Addgene’s collection. Green fluorescent aptamers Spinach...s group created Spinach2 using site-directed mutagenesis of Spinach. In doing so, they identified the ...

...an unpublished set of Zika gene plasmids to Addgene’s repository. The genes, from the Zika virus Asian...vectors targeting their genes of interest to generate mAID fusions to endogenous genes. Cell lines with the...targeting a total of 20,121 genes within the human genome (the identity of the genes and target sequences can...expression level for their gene of interest. Although the parts involved in altering gene expression are relatively...promoters and 140 genes, including constructs with unique tissue-specific promoters expressing GFP, tetracycline...contains ~90,000 gRNAs that target ~15,000 genes with 6 gRNAs per gene. The second is a "supplementary" library...library to generate a population of HEK293FT cells expressing a single gRNA targeting a single gene. The gRNA...

...Additional Resources on the Addgene Blog Treating Muscular Dystrophy with CRISPR Gene Editing Learn to Use ...activated HIV-1, measured via HIV-1 promoter-driven GFP expression. In cell lines that can produce HIV-1 ...Cas9 have also been used to create mutations in genes essential to HIV-1 replication. However, this approach...modification targeting either the LTRs or essential genes. Many of these escape mutations were located near...Genomes from Human T-lymphoid Cells by CRISPR/Cas9 Gene Editing. Sci Rep. 2016 Mar 4;6:22555. PubMed PMID...Screens Additional Resources onf Addgene.org Check out Addgene’s newly updated CRISPR Guide! Find CRISPR...Activation Mediators (SAM) system available from Addgene. This system uses the dCas9-VP64 fusion commonly...

...may be the hot new way to manipulate gene expression, but other gene manipulation systems remain valuable...important ways to spatially and temporally control gene expression, especially in in vivo models, and new...-2.1 construct can express one of four colors (n-GFP, RFP, YFP or M-CFP.) The construct contains two tandem...mOrange2, jellyfish EGFP, and sea anemone mKate2.) They then successfully generated custom antibodies to...-2. The general structure of Brainbow-1.0 is retained in Brainbow-3.0, but with mOrange2, EGFP and mKate2...at least six generations. In addition to the neuronal pThy1-Brainbow constructs, Addgene also has two ...Brainbow plasmids available from Addgene! Find Brainbow Plasmids @Addgene: Brainbow 1.0, 1.1, 2.1 plasmids...

...Figure 1: MTRIA biosensors are generated by inserting a circularly-permuted GFP into an intracellular loop...new to Addgene this year. Feel the love with MTRIA sensors One of the latest additions to Addgene’s catalog...imaging. Vladislav Verkhusha’s lab has generated a new genetically encoded calcium indicator using small...on the Addgene blog Fluorescent Proteins 101: Fluorescent Biosensors Resources on Addgene.org Fluorescent...Here at Addgene’s headquarters, the skies are getting dark and wintry with the end of year approaching...localized for synaptic glutamate response. They generated variants optimized for different applications ...St-Pierre lab and colleagues have developed a new genetically-encoded voltage indicator (GEVI) called JEDI-...

... transcription of a target gene, it therefore determines the timing of gene expression and largely defines...These permit exquisite control of gene expression and tracing of gene expression across development. One...of herpes simplex virus immediate early gene expression. Genes Dev. 1988 Jun;2(6):718-29. doi: 10.1101...systems are genetic tools that consist of two parts, both of which must be present for a gene of interest...placing UAS upstream of a transgene permits directed expression of that gene in cells that also express...expression of reporter genes, or to combine reporter expression from one promoter with transgene or siRNA expression...together for complex genetic analysis in Drosophila or for precise control over multiple genes in synthetic systems...

...µg/mL Anti-GFP [N86/38.1R] (Addgene #180084), 0.01 µg/mL Anti-HA [12CA5] - Chimeric (Addgene #198003)) ... Strong expression: A CMV promoter drives robust gene expression in mammalian cells. Flexible tagging:...tag. Selectable and stable: A puromycin resistance gene makes stable cell line creation a breeze. Modular...labeling By Alyssa Shepard To fully understand how genes are regulated, it's important to know what proteins...identify both new and known regulators at the FOS gene promoter and the MYC locus (Figure 5). Figure...identification of heat-shock proteins at the FOS gene promoter using TurboCas, streptavidin immunoprecipitation...very quick and efficient way to learn more about gene regulation. Find the TurboCas plasmid here! Cenik...

... bright enough to detect endogenous tagged genes generated through CRISPR/Cas9 knock-in. Finally, NLuc...Fusion Protein, codon-optimized Firefly luciferase genes (e.g., luc2), and NanoLuc® Luciferase are available...fluorescent protein that could be co-imaged with GFP using the same wavelength for excitation. The result...such as the dynamics of subcellular structures and gene expression. You must see the movies demonstrating...al. (2016) CRISPaint allows modular base-specific gene tagging using a ligase-4-dependent mechanism. Nature...when employing optogenetic regulators to initiate the event being monitored. Optogenetic regulators respond...wavelengths overlap with optogenetic initiation wavelengths, excitation of the optogenetic regulator leads to...

...protein, permeabilized cells  Cy5NTAc 32 25-45 GFP (fluorescein is similar)   Lumio Red Tc: CCPGCC 48...in setting up a FRET-based assay or screen (the Addgene blog has an excellent primer on FRET and how to...Fluorescent proteins (FPs) are the gold standard for genetically-encoded FRET donors (Fig. 1). Scores of FP variants...presented (Fig. 1) which can be carried out using Addgene plasmids. His-tag labeling reagents You may already...Gradia Lab has deposited a gateway vector with Addgene, where you can insert your protein of interest,...PubMed Central PMCID: PMC4380784. Resources on the Addgene Blog Get Tips for Using FRET in Your Experiments...Your Fluorescent Protein Basics Resources on Addgene.org Find Plasmids for FRET Browse Empty Plasmids...

...labeling. Size is also a factor. Bulkier tags, such as GFP, may interfere with protein expression or function...the dye and the endogenous target, and thus no genetically encoded tag is required. These dyes can also ...be SNAP-tag® and HaloTag®. These consist of a genetically encoded enzyme variant tag that covalently reacts...maintained through mechanisms like fission, fusion, biogenesis, and mitophagy, can be tracked using dyes like...Keller, P. J., Ji, N., & Lavis, L. D. (2017). A general method to fine-tune fluorophores for live-cell ....1016/j.bbamem.2013.09.005 More resources from Addgene Fluorescent protein plasmids and resourcesA practical...

...measurable through an epitope tag such as FLAG/His/GFP. Fluorescent protein tags, which correlate closely...procedure and application to the major ALS/FTD disease gene C9ORF72. eLife, 8, e48363. https://doi.org/10.7554...101: Validation Resources on addgene.org Addgene's Antibody Guide Addgene's Antibody Protocols... outlined below. Knockout cell line approach Genetic knockout-based validation can conclusively prove...you are working with cells that are amenable to genetic knockout, this represents a very robust approach...to start with a screening procedure in easily genetically modified cell types (top). The specific approach...10.1038/s41598-019-55133-7 More resources from the Addgene blog Introduction to Gating in Flow Cytometry Antibodies...

...reprogramming transgenes and the up-regulation of corresponding endogenous pluripotency genes. Retroviral...disrupt the expression of tumour suppressor genes and/or oncogenes, potentially leading to the presentation... Shao, L., & Wu, W. (2010). Gene-delivery systems for iPS cell generation. Expert opinion on biological...a single polycistronic vector or multiple single-gene vectors can be used to package reprogramming factors... the transposase. The donor plasmid contains the gene(s) of interest flanked by terminal repeat sequences...cassette comprised of OCT4, SOX2, LIN28, NANOG, and a GFP reporter. Minicircles are removed from cells by dilution...demethylation and expression of endogenous pluripotency genes (i.e. Oct4) are other measures of reprogramming....

...understanding of B. subtilis gene functions and interactions. The single-gene deletion library is derived...studying the phenotype of an essential gene, the CRISPRi Essential gene Knockdown library was constructed ...Bacillus subtilis libraries with Addgene – the B. subtilis Single Gene Deletion Library—Kanamycin and the...compatible cloning sites for easy cloning of your genes of interest. In order to improve the study of specific...cassette. However, it was unknown whether variations in gene expression from these polycistronic cassettes could... stoichiometry impacted reprogramming and global gene expression patterns. Polycistronic cassettes that...-positive assay) as well as stabilization (Nanog-GFP reporter activation and silencing of factor-linked...

... this gene. Should you cut it at the beginning of the coding sequence to delete the entire gene? Perhaps... in vitro, and using it in mouse embryos to generate gene modified mouse lines. Hopefully this post will...they have been generated (more on that later). Find Validated gRNAs to Target Mouse Genes To begin the ...chances of gene modification in vivo. Find Plasmids for Testing Your gRNAs in vitro at Addgene CRISPR/Cas9...decide to do a gene modification experiment in mice to the day you have your F0 generation (barring life...Haoyi, et al. "One-step generation of mice carrying mutations in multiple genes by CRISPR/Cas-mediated ...blogger Samantha Young. The use of CRISPR/Cas9 for gene editing has expanded since its adaptation for use...

...neurotransmission. But with approximately 950 different human genes encoding GPCRs (Takeda S et al., 2002) and many ...2002) Identification of G protein-coupled receptor genes from the human genome sequence. FEBS Letters 520... acceptor GFP2-Gγ chimeras, and two untagged Gβ constructs. For the Gγ constructs, the GFP2 tag is fused... Gα-Rluc8 and Gγ-GFP2 will be in close association, resulting in a high level of GFP2 emission. Following... exhaustive refinement, it was not possible to generate an optimal Rluc8-Gα construct for two of the 16...interest, a Gα-Rluc8 plasmid, and the appropriate Gγ-GFP2 and Gβ constructs. Cells are then treated with the...complex will disassociate, resulting in a decreased GFP2 emission. The activation state can be measured by...

...6 letter representation of the gene (or DNA sequence). pBACKBONE-hGene Tip: Often researchers will add...pBACKBONE-Flag-hGene(Q295A)-EGFP  Wildtype version: pBACKBONE-Flag-hGene-EGFP or pBACKBONE-Flag-hGene(wt)-EGFP...would list it first. pBACKBONE-Flag-hGene If there was also an EGFP fused to the C-terminal of your insert...would list it after the insert. pBACKBONE-Flag-hGene-EGFP Other considerations  If your insert contains...be included in the plasmid name. Mutations are generally listed as the amino acid change and not a nucleotide...

...magnetic responses: EGFP, mScarlet, and AsLOV2. After several rounds of semi-random mutagenesis and screening.../nn.4265.  Additional resources on addgene.org Find Optogenetics Plasmids Find Biosensor Plasmids Visit... al., 2024). They deposited their plasmid with Addgene, hoping to inspire others to explore the phenomenon...Although a wide range of fluorescent proteins and optogenetic tools exist, humans (like many organisms) are...magnetoresponsive fluorescent protein. Controlling an optogenetic or fluorescent tool with magnets would enable... molecules like proteins with magnetic fields generally appears to be weak — if they exist at all. Andrew...protein they tested — E. coli expressing good ol’ EGFP — showed a slight but detectable change in fluorescence...

...comparing gene activation when enhancers were active or silent, they could identify the target genes of previously...efficient. In general, epigenetic chromatin modifications are correlated with alterations in gene expression...updated Jun 24, 2020. Epigenetic modifications are an additional layer of control over gene expression that ...proteins that help package DNA, were associated with gene expression or silencing. To alter DNA modifications...oxidation and repair factors that allow for increased gene activation at the targeted site compared to other...the only CRISPR-based technology designed to alter gene expression. Fusing dCas9 to a transcriptional activator...a CRISPR-Cas9-based acetyltransferase activates genes from promoters and enhancers. Nature Biotechnology...

...include: Optogenetics AAV Control AAV Biosensor AAV And more! Optogenetics Optogenetics continues ...Nuc-flox(mCherry)-EGFP 27056 AAVrg pAAV-Ef1a-DIO EYFP 50457 AAV2 pAAV-hSyn-DIO-EGFP   Biosensor AAV...pOTTC1032 - pAAV EF1a Nuc-flox(mCherry)-EGFP 50457 AAV2 pAAV-hSyn-DIO-EGFP Recombinases Plasmid Serotype...viral vectors 101 eBook Resources on Addgene.org Learn about Addgene's viral service Find viral protocols...you pick the best AAV for your experiment. The Addgene viral service has now expanded to offer many of... Plasmid Serotype Name 51502 AAV5 pCAG-FLEX-EGFP-WPRE 114471 AAV1, AAV5 pAAV-Ef1a-fDIO mCherry ...pEF1a-DIO-FLPo-WPRE-hGHpA    Additional resources on the Addgene blog Read all of our viral vectors blog posts ...

...AAV9 Optogenetics Adesnik New viral vector pAAV-nEF-Con/Foff DREADD Gi-mCherry AAV8 Chemogenetics Deisseroth... pENN.AAV.EF1a.eGFP.WPRE.rBG AAV PHP.eB Controls Wilson New serotype pAAV-CaMKIIa-EGFP AAV PHP.eB Controls...latest in Addgene's viral vectors services and resources!   More resources on the Addgene blog Addgene's...What's new in Addgene's ready-to-use viral vectors repository? Quite a bit! In this post, we'll share...Table 1: New ready-to-use viral vectors in the Addgene repository  Name Serotype Category Depositor...Deisseroth New viral vector pAAV.CMV.PI.EGFP.WPRE.bGH  AAV PHP.eB Controls Wilson New viral vector ...the full viral services collection or check out Addgene's Data Hub to view viral vector reports from other...

...victoria, AvGFP forms a homodimer. In corals, the red fluorescent proteins form tetramers. In general, fluorescent... develops genetically encoded FRET based biosensors.     Additional Resources on the Addgene Blog Which...detect weak dimerization tendency, as is present in EGFP. Constantini et al, (2012) developed a cell-based... properties of new fusions can be compared with mEGFP or mTurquoise2, which are established true monomeric...Amsterdam). He develops, characterizes and uses genetically encoded fluorescent probes. You can follow him... Protein Biosensors Additional Resources on Addgene.org Find Empty Backbones with Fluorescent Tags Find...

...for doing gene targeting, reporter generation, controllable gene knockouts, and inducible gene inactivation...Alec Kimmelman targets mouse metabolic genes with ~6 gRNAs per gene.  CRISPR-Act3.0, developed by Yiping... Shirley Liu's lab targets all mouse genes with 10 gRNAs per gene. The guides are optimized to maximize... to develop CRISPR-based approaches for studying gene function in these systems. As a part of this effort...Yiping Qi's lab, allows simultaneous gene activation in plants.  FnCas12a, unlike other Cas12a nucleases identified...opportunities for investigating genetic mechanisms related to morphogenesis, cell fate specification, and...Lab have compiled a new toolkit for performing genetics studies in organoids. The Organoid EasyTag System...

...provide the Brunello (targeting human genes) and Brie (targeting mouse genes) pooled libraries from the labs...targeting EGFP, BRAF, and MAP3K4. The gRNAs targeting EGFP can be used as positive controls for gene editing...editing. You can also enhance expression of your gene of interest with a variety of Cas9 activators available...editing or gene activation, while the BRAF and MAP3K4 gRNAs allow you to target these widely studied signalling...pooled gRNA libraries that can be used to screen for genes involved in a wide variety of cellular processes...80,000 gRNAs to target around 20,000 individual genes. We provide enough lentivirus for you to both perform...and member of the Addgene Advisory Board) says: "When you're setting up to do a genetic screen, you need...

...promoter. Use this for doxycycline inducible (tet-off) gene expression! We've also added AAV1 and AAV5 preparations...variants of a new optogenetic tool called PdCO, deposited by the Yizhar lab. Addgene is now offering AAV1... pAAV_hSyn-SIO-stChrimsonR-EGFP-P2A-PdCO-miniWPRE AAV1, AAV5 Optogenetics Yizhar New viral service with...AAV1, AAV5 Optogenetics Yizhar New viral service with multiple serotypes pAAV_hSyn-PdCO-EGFP-WPRE AAV1... Brian O'Neill.  More resources on the Addgene blog Addgene's Data Hub What is a Virus (and a Viral Vector...What's new in Addgene's ready-to-use viral vectors repository? Quite a bit! In this post, we'll share...to our viral vectors repository since March.  Addgene is now offering an AAV2 CAG-driven tTA expresser...

...exhibiting its versatile utility in quantifying multiple gene editing activities. Click here to find the ACE ...Wanisch  Listen to the optogenetics podcast segment LOVTRAP is an optogenetic system that was recently... the University of North Carolina (plasmids at Addgene). It consists of two domains, LOV2 and Zdk1, where...function is more specific and much faster than any genetic or pharmacological manipulations on cells, and ... riboswitches used in the Riboglow platform at Addgene. Braselmann E, et al. Nature Chemical Biology...real-time. The reporter constitutively expresses eGFP to indicate reporter abundance, and mCherry fluorescence...involves tracking the ratio of mCherry-positive to eGFP positive cells through fluorescence microscopy or...

...ampicillin resistance gene, you’ll find the full name of the gene, the name of its gene product, and a description... plasmid features such as antibiotic resistance genes, selectable markers, promoters, origins, tags, and... Overview of SnapGene powered plasmid maps at Addgene  Three factors make our SnapGene powered plasmid...powered by GSL Biotech's SnapGene Server Software. With the backing of SnapGene’s sequence viewer software...the features you need quickly. SnapGene’s Feature Library - SnapGene’s software seamlessly identifies ...nucleotides, such as the fluorescent proteins EGFP and mEGFP. Many of these features, such as Cas9, weren...scientists) for suggesting these changes, and the SnapGene and Addgene software development teams for working tirelessly...

...been used for many years for in vivo delivery and gene therapy and represent one of the most studied viruses...CRISPR-Cas. Figure 1: Schematic of targeted gene knock-out (left) and targeted DNA knock-in (right...binds to target DNA. In the final step of targeted gene knock-out (left), a fragment is deleted from two... This capacity is large enough to carry the Cas9 gene and a gRNA expression cassette in one single viral...Joining (NHEJ) DNA repair mechanism to knock out genes as a result of an insertion, a deletion or a frameshift...with Cas9 and a gRNA designed to target a mutated gene. In a Scientific Reports paper introducing the delivery... CRISPR/Cas9-derived RNA-guided nuclease-induced gene disruption frequencies in the various cell types...

...could replace one of the reporter genes above with your gene or genes of interest. Some of the plasmids...reporter gene). Addgene's collection offers a variety of plasmids to express two or more genes, some of... that are expressing your gene, especially if you are studying a novel gene, is not always a straightforward... your gene of interest, scientists have instead developed novel methods to co-express your gene along ...with a reporter, such as a fluorescence gene or a resistance gene. These reporters allow you to easily screen...more of the genes replaced.  Additionally, 2A peptides could be PCR-cloned between your genes of interest...more than two genes. Plasmid Name Multicistronic Element Expression Type MSCV-IRES-EGFP IRES Retroviral...

... what part of a gene enhancers like to interact with, and how many promoters or genes each enhancer is...plot line in gene regulation. When chromatin conformation data is compared with changes in gene expression...dynamic role in gene regulation The high resolution and multiplexed chromatin ‘photos’ generated with CAPTURE...changes in gene expression and determine who’s a key driver of these changes. By generating a more vivid...updated Jul 2, 2020. Promoters may be the star of gene regulation, but enhancers and chromatin looping ... bound by transcription factors, can increase a gene’s transcription. Sometimes enhancers are located ...located thousands of base pairs away from the gene they regulate, but are brought in proximity by the looping...

...luciferase or eGFP) flanked by the EBOV-specific gene start and gene end sequences. Thus, the minigenome...replication and viral gene transcription (6). The EBOV genome contains seven viral genes, each flanked by ...sequence (gene start) and a transcription termination/polyadenylation signal sequence (gene end). A minigenome...Figure 1). All viral genes are removed and replaced by a single (non-viral) reporter gene (e.g. firefly luciferase...Cloning the viral genes in this plasmid would be more difficult compared to if each gene were encoded in...transcribe the reporter gene (e.g. firefly luciferase). The polymerase recognizes the gene start signal and ...of the reporter gene. The polymerase will elongate the mRNA until it reaches the gene end signal, where...

...observed in mammalian cells is to tag an endogenous gene. By imaging cells (or tissues) that produce an endogenous...fluorescent proteins or using endogenous tagged genes) can be used to verify performance under different... develops genetically encoded FRET based biosensors.     Additional Resources on the Addgene Blog Which... QY (EC*QY, sometimes normalized to the value of EGFP). The higher the number, the higher the theoretical...mVenus as a reference protein is available from Addgene. Another way to dodge cellular variation observed...Amsterdam). He develops, characterizes and uses genetically encoded fluorescent probes. You can follow him... Protein Biosensors Additional Resources on Addgene.org Find Empty Backbones with Fluorescent Tags Find...

... Figure 1: In conventional gene targeting (A), you must first generate your desired mutation in mouse...mutations, the 8-10 month process of conventional gene targeting is lowered to about 2-3 months. But certain...example of inserting a 2A-FlpO cassette into the Fgf8 gene. Miura et al. (2018) suggest using a 50-80 bp sequence... mouse model generation easi-er! Generating mice before CRISPR The process of generating knock-in mice...process of mouse model generation simpler, as described in an earlier post on generating CRISPR mouse models...after editing. How to use Addgene plasmids for Easi-CRISPR You can use Addgene plasmids to create a repair...template. Figure 2: Overview of ivTRT to generate an ssDNA repair template. Although effective...

...excellent series of posts by Joachim Goedhart on the Addgene blog – outlining various important considerations...relating to that specific protein (for instance: EGFP).  Users can “favorite” certain proteins, or create...github @tlambert03.   Additional Resources on the Addgene Blog Fluorescence Microscopy Techniques - Which...Proteins for Multicolor Imaging Resources on Addgene.org Browse all fluorescent protein plasmids Find...

...Entrez Gene matches to your insert. Select the appropriate gene if it appears in the Entrez Gene list, ...correct Entrez Gene does not appear in the auto-populated list, try entering the gene's Official Full ...known as" aliases in the Gene/Insert box. Once you have selected the appropriate gene from the list, save ...bacterial strain(s) through Addgene. Description – please indicate the gene cloned into your plasmid, any... for the gene or insert. Step 4: Growth and Distribution Please indicate which strain Addgene should propagate... Encodes insert(s) A plasmid with one or more genes, protein coding sequences, or other elements inserted...Empty backbone A plasmid designed for cloning a gene/insert into it. Empty backbones often contain epitope...

...in neurons. (b) Genetic cargo can be customized to control transgene expression. Gene regulatory elements... relatively even distribution of target gene expression (EGFP, in the depositing manuscript) across the....     Figure 1: AAV toolkit for targeted gene delivery to the brain. The AAV (a) capsid, (b) cargo...Abbreviations: AAV, adeno-associated virus; GRE, gene regulatory element; NHP, non-human primate. ...Systemic AAV vectors for widespread and targeted gene delivery in rodents. Nature Protocols;14, 379–414...2017). Engineered AAVs for efficient noninvasive gene delivery to the central and peripheral nervous systems...Gradinaru, V. (2022). Engineered AAVs for non-invasive gene delivery to rodent and non-human primate nervous...