... for a researcher to come into contact with than GFP. Similarly, if the viral vector carries an shRNA ...viruses grown in the lab are gamma retrovirus, lentivirus, adenovirus, and adeno-associated virus (AAV)...humans, earning this virus a BSL-2 classification. Lentivirus (a type of retrovirus) and gamma retrovirus require...and required in BSL-2 labs. When working with lentivirus, many organizations require or recommend disposable...you have to work with BSL-2 materials, such as lentivirus. Most of this work will need to be done in a ...that particles are not sprayed into the air. For lentivirus and other BSL-2 materials, you’ll need to use...for extra security. Finally, when spinning down lentivirus, you’ll need to use centrifuge bin covers, which...

...another marker, such as GFP. A researcher can use FACS to sort cells expressing GFP and later expand these...produce lentivirus? 293T cells are usually used to produce lentivirus. What dictates lentiviral host cell... Guides Lentiviral Guide Lentiviral Guide...FAQ References Glossary Lentiviral Components To increase the safety of lentivirus, the components necessary... below. Third Generation Lentiviral Plasmids 2nd vs 3rd Generation Lentiviral Systems Feature 2nd Generation...generation lentiviral systems? For a full description of 2nd and 3rd generation lentiviruses, please review...What is the difference between a lentivirus and a retrovirus? Lentiviruses are a subtype of retrovirus. From...

... later, Cre-dependent GFP expression was detected with direct fluorescence. GFP was not detected in the...viral genomes (vg)/cell, pAAV-CAG-FLEX-rc [Jaws-KGC-GFP-ER2] (Addgene 84445-AAVrg) alone at 1.1E6 vg/mL, ... prep # 55632-AAVrg). pAAV-CAG-FLEX-rc [Jaws-KGC-GFP-ER2] was a gift from Edward Boyden (Addgene viral...AAV vectors collection. AAV Data Hub Lentivirus Production Lentiviral preparations are produced in 293T ...performed on lentiviral preparations that have been stored at -80 °C and thawed. Lentivirus titration is...for in our reported titers. Lentiviral vectors are titered using a lentiviral droplet digital PCR (ddPCR...viral production methods and quality control for lentivirus and AAV. Viral...

...the viral protease, Nsp5. Our set also includes a GFP construct as a control. For easier portability, all...vector background, so you can package them into lentiviral vectors if you need to. Additionally, they have...

...Addgene has several great options: pLV hUbC-dCas9-T2A-GFP for removable dCas9 expression and pLM-CMV-R-Cre....One good system to balance these needs is the lentiviral-based expression of floxed dCas9 so that it may... stable gRNA expression can be achieved using lentiviral backbones expressing blasticidin or puromycin...

...cells in vivo, the non-immunogenic violet-excited GFP variant Vex. To understand the impact of this gene...high-throughput manner. They designed the pXPR_053 lentiviral expression vector, which contains an sgRNA insert...

...activator with 2A GFP 61423 : Expresses the MS2-P65-HSF1 activator helper complex with 2A GFP 61424 : sgRNA...(SapI)_hSyn-GFP-KASH-bGH (SpGuide acceptor). This is a AAV plasmid for sgRNA cloning. GFP-KASH fusion ...PX552; U6 promoter-driven; for sgRNA cloning; has GFP-KASH for FACS sorting SaCas9 + single guide RNA: ...system - lentiCRISPR - sgRNA and SpCas9 together 2 vector system - lentiCas9-Blast and lentiGuide-Puro (...#61425 - #61427 are intended for lentivirus production : 61425 : lentiviral vector encoding dCAS9-VP64 with... recombinase-2A-EGFP-KASH 60231 : sgRNA cloning backbone with Cre recombinase-2A-EGFP-KASH Detailed backbone...recombinase-2A-EGFP-KASH and an sgRNA. #60231 - AAV:ITR-U6-sgRNA(backbone)-hSyn-Cre-2A-EGFP-KASH-WPRE-shortPA-ITR...

..., Hess et al. successfully evolved wild type GFP to EGFP using CRISPR-X and subsequent FACS sorting. They...with unique tissue-specific promoters expressing GFP, tetracycline response elements, and shRNAs many ... of DsRed connected to a pH-sensitive variant of GFP (SEP) by a 9 amino-acid linker (see Figure 1). The...the authors packaged it into lentivirus and used the resulting lentiviral library to generate a population...Science. 2013. PubMed PMID: 23328393 Wu lab human lentiviral CRISPR library     CRISPR pooled libraries enable...applications. First, GMAP-compatible backbones for lentivirus LV 1-5 and retrovirus RV 2-5 were constructed...Library. This complex second-generation CRISPR lentiviral library targets nearly all human protein-coding...

..., Myc and SV40 Nuclear Localization Signals, and GFP. B) Workflow to assess Cas9-PAGE editing with viral...facilitates endosomal escape. Combining Cas9-PAGE with lentiviral sgRNA expression yielded editing of 80–90% in...

...check out our article: Fluorescent Proteins 101: GFP Fusion Proteins - Making the Right Connection. Cellular...theoretically possible, but likely more challenging, with lentiviral expression. For more detail on how to design ...

...Vectors Read about Reporter Gene like Luciferase and GFP Resources on Addgene.org Find Plasmids for Your... as RNAP II promoters is commonly employed in lentiviral and retroviral constructs and we will discuss...

...activated HIV-1, measured via HIV-1 promoter-driven GFP expression. In cell lines that can produce HIV-1 ...people worldwide are currently infected with the lentivirus HIV-1. Today, HIV-1 can be controlled with antiviral... with HIV-1. For two HIV-1 strains, Cas9/gRNA lentiviral expression significantly reduced the HIV-1 copy...PubMed Central PMCID: PMC4778041. Find CRISPR and lentiviral packaging plasmids used in this publication at...

... includes things like viral vectors that deliver GFP.   Figure 3: A) Cumulative AAV preps available...2016). Practical Considerations for Using Pooled Lentiviral CRISPR Libraries. Current Protocols in Molecular...

...vector Retroviral vectors: neomycin (pSIR-neo) , GFP (pSIR-GFP) , DsRed (pSIR-DsRed-Express2) , human CD2 (...amplification 1 vector system: lentiCRISPR v2 2 vector system: lentiCas9-Blast and lentiGuide-Puro packaging plasmids...sgRNA cloning lentiCRISPR v2 packaging plasmids: pVSVg , psPAX2 ; positive control: CMV-EGFP PDF 2.3 MB ...Musunuru CRISPRs in human pluripotent stem cells pCas9_GFP ; gRNA empty vector Link O'Connor-Giles Fly: gRNA...plasmids: pVSVg , psPAX2 positive control: CMV-EGFP Kits are also available (mouse or human libraries) PDF...

...typically used for gRNA May contain reporter gene (e.g. GFP) to identify and enrich positive cells, or selection...transfer vectors May contain reporter gene (e.g. GFP) or selection marker to identify and enrich positive...components to make lentiviral particles (for details about lentivirus, see our Lentivirus Guide ) Stable,...high efficiency Lentiviral transduction Cas9 and gRNA can be present in a single lentiviral transfer vector...

...Calcium erGAP3 (GFP-Aequorin Protein) for imaging of Ca+ dynamics in endoplasmic reticulum GFP-Aequorin Protein...FlincG3 (GFP-based cGMP sensor) for imaging in C. elegans neurons Using a Robust and Sensitive GFP-Based ...2020 GENIE Project Calcium jGCaMP7 High-performance GFP-based calcium indicators (Constitutive or Cre-dependent...cyclic AMP) Signaling reporter island (SiRI) with GFP-based fluorescent reporter cAMPr Spatial Multiplexing... FLAMP Plasmids Jun Chu cGMP (cyclic GMP) FlincG GFP-based cGMP sensor Differential patterning of cGMP... vascular smooth muscle cells revealed by single GFP-linked biosensors. Proc Natl Acad Sci U S A. 2008...autophagic flux by pH-sensitive fluorescence (pMRX-IP-GFP-LC3-RFP) An Autophagic Flux Probe that Releases an...

...cassette comprised of OCT4, SOX2, LIN28, NANOG, and a GFP reporter. Minicircles are removed from cells by dilution...considered safe for clinical applications.     Lentiviral: Lentiviral vectors infect dividing and nondividing...Excisable Cre-lox lentiviral vectors: This approach is the same as using a standard lentiviral vector, but ...transgene reactivation, residual expression Lentivirus Integrating High Low Efficient and Stable Genome...iPSC-transplanted mice. Cre-deletable or Tet-inducible lentiviruses address some of these concerns, but overall ...reprogramming factors driven by one promoter. While lentivirus has similar rates of reprogramming when compared...available from Addgene. Another drawback is that lentiviral vectors insert in different genetic locations...

...typically used for gRNA May contain reporter gene (e.g. GFP) to identify and enrich positive cells, or selection...transfer vectors May contain reporter gene (e.g. GFP) or selection marker to identify and enrich positive...components to make lentiviral particles (for details about lentivirus, see our Lentivirus Guide ) Stable,...high efficiency Lentiviral transduction Cas9 and gRNA can be present in a single lentiviral transfer vector...

...Cre-dependent expression of diphtheria toxin receptor (DTR)–GFP fusion protein. Azim et al. Nature. 2014 Apr 17. ... Proc Natl Acad Sci U S A. 2023 Aug 22. Use a lentiviral FRET-based reporter to study tau seeding in mammalian...

...tools. pLV hUbC-dCas9-MQ1(Q147L)-EGFP is also available for lentiviral transduction. Lysine-specific Demethylase...Core, as well as pLV-dCas9-p300-P2A-PuroR for lentiviral expression. Figure 1: dCas9-p300 adds ...decrease methylation and activate transcription. A lentiviral vector with the same modifier, Fuw-dCas9-Tet1CD...Jaenisch’s lab in plasmid form or as ready-to-use lentivirus. Figure 2: dCas9-Tet1 demethylates cytosines...) Vlatka Zoldoš’ lab has deposited pdCas9-DNMT3A-EGFP and pdCas9-DNMT3A-PuroR for targeted cytosine methylation...methylation in mammalian cells. Co-expression markers EGFP and PuroR enable sorting and selection of transduced...Tet-dependent (TetO-dCas9-D3A) constructs. For lentiviral expression, Fuw-dCas9-Dnmt3a and Fuw-dCas9-Dnmt3a-P2A-tagBFP...

...of producing CRISPR lentivirus. As we’ve heard from scientists, producing lentivirus is not always an easy... can make lentivirus production frustrating - a researcher can complete an entire lentiviral production...titering, testing, and providing lentiviral preps of select lentiviral vectors from the repository for ... in lentiviral format. Finally, while we can’t currently produce custom gRNA-containing lentivirus, we...providing lentiviral preps containing highly requested gRNAs including those targeting EGFP, BRAF, and... make new lentiviral preps on your own, fear not - anytime you order ready-to-use lentivirus from Addgene...you can use along with lentiviral packaging plasmids to create your own lentivirus stocks. Our research ...

...for inducible CRISPR inhibition (CRISPRi). A) Lentiviral vector design. B) Strategy for leak-free transcriptional...circularly permuted green fluorescent protein or cpGFP) to the petunia SL enzymatic receptor DAD2. SLs ...which modulates the fluorescence intensity of the cpGFP without altering that of the internal control (Fig...highlights from the past couple months: A new lentiviral CRISPR library from Alec Kimmelman targets mouse...

...from three to two. Lentiviral delivery of BioTAP-tagged dCas9 and gRNAs. Using lentiviral delivery of the...more gRNAs Two separate lentiviruses delivering 1) a BioTAP-tagged dCas9 and an eGFP tag, and 2) gRNAs ...

...Multicistronic Element Expression Type MSCV-IRES-EGFP IRES Retroviral pMSCV-pBabeMCS-IRES-RFP IRES Retroviral...MSCV-IRES-Luciferase IRES Retroviral pWPI IRES Lentiviral AmCyan-P2A-mCherry P2A Mammalian pC5Kan-P2A...-P2A P2A Insect pUltra P2A and T2A Lentiviral Ac5-STABLE2-neo T2A Insect   For the co-expression....) PubMed. 3. Highly Efficient Multicistronic Lentiviral Vectorswith Peptide 2A Sequences. Ibrahimi et...

...categories (e.g., for a lentiviral vector to be used in mammalian cells, check Lentiviral and Mammalian Expression...Please describe any tags or fusions (e.g. His, FLAG, EGFP, etc) on your insert. Use the drop-down menu to ...be prone to recombination in bacteria) such as lentiviral, retroviral, and AAV plasmids, we recommend the...

...deaminases like APOBEC. To achieve this they created a lentiviral vector system that monitors both APOBEC- and ...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...

...non-viral) reporter gene (e.g. firefly luciferase or eGFP) flanked by the EBOV-specific gene start and gene...incapable of producing infectious virus (similar to lentiviral vectors) because they do not encode all of the...Addgene.org Browse Our Viral Vector Plasmids Read Our Lentiviral Vector Guide Read Our AAV Guide  ...

...contain a reporter gene (for example, luciferase or GFP) that offers a read-out of the activity of the genetic...these plasmids to create viral particles, such as lentiviral, retroviral, AAV, or adenoviral particles, that...viral service offers select ready-made AAV and lentiviral particles. Visit our viral service page to learn...

...fluorescent marker like green fluorescent protein (GFP), creating a customizable DNA or RNA label for fluorescence...CRISPR libraries What Are Pooled Lentiviral CRISPR Libraries? Pooled lentiviral CRISPR libraries are a heterogeneous...need to generate lentivirus containing the entire library (Figure 8D). This lentivirus will then be used...genome-wide screens using CRISPR involves pooled lentiviral CRISPR libraries. Figure 8: Overview of genome-wide...heterogeneous population of lentiviral transfer vectors. In most cases, each plasmid contains an individual...Figure 8A), then cloned in a pooled format into lentiviral transfer vectors (Figure 8B). Libraries come ...library is suitable for your experiment. Although lentiviral libraries are the most popular method for CRISPR...

...fluorescent marker like green fluorescent protein (GFP), creating a customizable DNA or RNA label for fluorescence...CRISPR libraries What Are Pooled Lentiviral CRISPR Libraries? Pooled lentiviral CRISPR libraries are a heterogeneous...need to generate lentivirus containing the entire library (Figure 8D). This lentivirus will then be used...genome-wide screens using CRISPR involves pooled lentiviral CRISPR libraries. Figure 8: Overview of genome-wide...heterogeneous population of lentiviral transfer vectors. In most cases, each plasmid contains an individual...Figure 8A), then cloned in a pooled format into lentiviral transfer vectors (Figure 8B). Libraries come ...library is suitable for your experiment. Although lentiviral libraries are the most popular method for CRISPR...