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We narrowed to 224 results for: cas9 expression plasmid

Showing: 21 - 40 of 224 results

  1. 27 Hot Plasmids from 2016

    Type
    Blog Post
    ...Kit contains Cas9, Cas9 nickase, and Cas9 activator expressing plasmids that can co-express 1-8 gRNAs. ...lines expressing it. The Church lab fixed this issue by creating a new dCas9-VPR expression plasmid that...then transfected with a Cas9-expressing plasmid to generate a population of mutant cells. The mutant cells...the Liu lab’s own pAc-sgRNA-Cas9 insect expression backbone, which expresses the gRNA from a Drosophila...deposited their Cas9 and gRNA backbone plasmids with Addgene. Additional control plasmids for the TKO library...parasites that constitutively express Cas9 with a library of sgRNA expression vectors (containing ten guides...Neurosci 2016. PubMed PMID: 26950006 Plasmid ID Plasmid Name Plasmid Type 74309 pcDNA3.0-TRPV4-p2A-ferritin-p2A-mCherry...

  2. What's New in CRISPR - December 2019

    Type
    Blog Post
    ...CRISPR plasmids available at Addgene. We will still periodically focus on specific CRISPR plasmid tools...capsid-based bionanoparticle Transient expression of CRISPR/Cas9 in cells has advantages like reduced off-target...bionanoparticle system for packaging Cas9/gRNA ribonucleoproteins for transient expression. They replaced the tetraloop...CRISPR guide Find CRISPR plasmids by function Find recently deposited plasmids ...toolkit for epitope tagging Lentiviral capsid-based Cas9/gRNA ribonucleoprotein delivery Cytosine deaminase...of bacteria that can cause disease in humans. Expression of this system is tunable via a rhamnose-inducible... a catalytically impaired Streptococcus pyogenes Cas9. They called this editor Target-ACE (adenine and...

  3. Hot Plasmids - February 2022

    Type
    Blog Post
    ...editing within a single expression cassette is advantageous to Cas9’s larger expression requirements (Figure...individually expressed proteins run in separate lanes. B: individually expressed and coexpressed proteins ...subset of the new plasmids and viral preps in the repository through our hot plasmids articles. These articles...recent plasmid deposits and we hope they'll make it easier for you to find and use the plasmids you need...Image from Santilli, et al. 2021 Plasmids are used to express 10, 15, 20, 30, 40, 50, 60, 80 and 100...through two polycistronic expression vectors (3570 lanes/50 mLs) coexpressing a set of either 10, 30, 50...need. If you'd ever like to write about a recent plasmid deposit please sign up here.  Here's what you'...

  4. CRISPR Methods for Bacteria: Genome Engineering, CRISPRa, CRISPRi, Base Editing, and More

    Type
    Blog Post
    ...available from Addgene consists of two plasmids: pCas9: carries Cas9 and chloramphenicol resistance pCRISPR...technology (Ronda et al., 2016). pMA7CR_2.0 expresses lambda Red and Cas9, which are separately inducible by L-arabinose.... citrea scarless editing plasmids Sheng Yang’s lab describes a two-plasmid system that combines recombineering...pCRISPomyces-1 includes Cas9, a tracrRNA, and a CRISPR array, while pCRISPomyces-2 contains Cas9 and a gRNA cassette...functions more similarly to CRISPR/Cas9 systems adapted for eukaryotes in that Cas9-mediated cleavage induces ...Tet responsive promoter to drive expression of a catalytically dead Cas9 enzyme allows reversible repression...knockdown expression of multiple target transcripts simultaneously. Marraffini lab's CRISPRi/a plasmids In addition...

  5. Your Top Requested Plasmid in 2016!

    Type
    Blog Post
    ...cell types. Purpose  SpCas9 This plasmid expresses Streptococcus pyogenes (Sp) Cas9 from a Cbh promoter...the top requested plasmid from 2016 is an expression vector for CRISPR nuclease, SpCas9 from the Zhang Lab...a versatile mammalian expression vector that you can use to express both SpCas9 and your gRNA in a variety...a versatile mammalian expression vector that you can use to express both SpCas9 and your gRNA of interest...sure, as is obvious from our most requested plasmid, SpCas9 is still going strong as the basis for many...without further ado, the most request plasmid in 2016 was... pSpCas9(BB)-2A-Puro (PX459) V2.0       Similar...sequence as directed by a gRNA. 2A-Puro In this plasmid, SpCas9 is fused to the puromycin resistance gene through...

  6. Targeting HIV-1 with CRISPR: Shock and Kill or Cut it Out?

    Type
    Blog Post
    ...or apoptosis with Cas9/gRNA expression. A pooled analysis found no evidence of Cas9 cleavage at predicted...promoter-driven GFP expression. In cell lines that can produce HIV-1 toxic proteins, CRISPR/Cas9 SAM caused apoptosis...target the HIV-1 5’ and 3’ LTRs and expressed them along with Cas9 in the T-cell line 2D10. PCR amplification...portion of the LTRs joined together. Clonal Cas9/gRNA-expressing cell populations were also immune to HIV...with HIV-1. For two HIV-1 strains, Cas9/gRNA lentiviral expression significantly reduced the HIV-1 copy...preventing re-infection would require sustained expression of Cas9/gRNAs, which could result in higher rates...were located near the Cas9 cut site, leading Wang et al. to conclude that some Cas9-derived indels may not...

  7. Plasmids 101: The protein expression toolbox

    Type
    Blog Post
    ...permanently induce expression? Check out our Retro expression vector and Lenti expression vector. Turn it...blog Plasmid 101: Inducible Promoters Plasmid 101: Repressible Promoters CRISPR 101: Which Cas9 Do I...turn off expression permanently is to knock out the gene for your protein of interest. CRISPR-Cas9 and a ...ablate protein expression. If you need the ability to control when you turn off expression for good, the...inducible expression (more on this below) will both achieve temporary repression of protein expression. Addgene... a gene (cDNA) can be done by either plasmid or virus. A plasmid carrying your cDNA can be targeted for...appropriate promoter to express your cDNA, depending on the level of expression you would like. Check out...

  8. What's New in CRISPR - March 2020

    Type
    Blog Post
    ...downregulate expression of specific Caulobacter genes using a catalytically dead Cas9 (dCas9) derived from...CRISPRa depended on continued expression of dCas9 and gRNAs whose expression varies depending on transgene...CRISPR plasmids available at Addgene. We will still periodically focus on specific CRISPR plasmid tools... lower average Cas9-independent off-target DNA editing, ~5–50-fold lower levels of Cas9-dependent off-...CRISPR guide Find CRISPR plasmids by function Find recently deposited plasmids ...fluorescent dCas9-KRAB and dCas9-VPR for quantifying and tracking cells that express both the dCas9 construct...repression of gene expression in Saccharomyces cerevisiae. The system consists of dCas9, the mammalian transcriptional...

  9. Four Base Editing Reporters to Monitor and Enrich Editing in Real-time

    Type
    Blog Post
    ...determined by comparing cells expressing only eGFP expression to cells expressing eGFP and mCherry. ACE was...single base editing by APOBEC-Cas9 Though the ACE reporter quantifies APOBEC-Cas9 efficiency in real-time, ...mCherry and a T2A site and expressed in cells. mCherry is constitutively expressed to identify successfully...efficiency of plasmid delivery to a cell. In contrast, TREE provides a read out for plasmid delivery and...integrated it into human and mouse cells that expressed either Cas9 or an optimized cytidine base editor. The...and enrichment of DNA editing by APOBEC–Cas9 or cleavage by Cas9 in living cells. Nucleic Acids Research... a APOBEC-Cas9n-UGI complex. So how exactly does this work? Figure 1: APOBEC- and Cas9-mediated ...

  10. Introducing an All-in-One CRISPR/Cas9 Vector System for Multiplex Genome Engineering

    Type
    Blog Post
    ...ease and efficiency for all-in-one CRISPR/Cas9 vectors expressing up to seven guide RNAs (gRNAs). Sakuma ...They went on to show that their CRISPR/Cas9-nuclease and CRISPR/Cas9-nickase vectors work successfully to...based on earlier evidence showing that a CRISPR/Cas9 plasmid could be injected directly into mice for genome...versions of Cas9, nuclease and nickase, but we are now adopting dCas9, dCas9-VP64, and dCas9-KRAB in our...all-in-one vector construction system for CRISPR/Cas9-mediated multiplex genome engineering is now available...one of the most advantageous properties of CRISPR/Cas9 compared to ZFNs and TALENs,” said Tetsushi Sakuma...Figure 1: Schematic overview of the all-in-one CRISPR/Cas9 vector construction system for multiplex genome ...

  11. Nanoblades: Tiny CRISPR Ninjas for Genome Editing Difficult Cells

    Type
    Blog Post
    ...compared to DNA transfection of gRNA and Cas9 expression plasmids. These results are in line with previous...releasing Cas9. Nanoblades can also deliver Cas9 variants, such as a transcriptional activator dCas9-VPR, ...Nanoblades by association with the the Cas9 portion of the Gag-Cas9 fusion protein. A single Nanoblade particle...along with the gRNA and Cas9. An “all-in-one” Nanoblades which carries a gRNA, Cas9, and a HDR donor template...centrifugation. Gag-Cas9 fusion protein The MLV Gag protein fused to a Flag-tagged SpCas9 is incorporated ...Gag in place of SpCas9. gRNA A gRNA targeting a DNA sequence of interest is expressed in 293T cells and...infiltrate cells. Options for CRISPR delivery include plasmid transfection, RNP electroporation, and viral transduction...

  12. Adenoviral Delivery of CRISPR/Cas9 Aims to Expand Genome Editing to Primary Cells

    Type
    Blog Post
    ...labs’ plasmids, including protocols, check out the plasmids at Addgene: pAdSh.PGK.Cas9 (expresses S. pyogenes...target cells. Within target cells, Cas9 machinery is transiently expressed and binds to target DNA. In the...capacity is large enough to carry the Cas9 gene and a gRNA expression cassette in one single viral particle...DNA Adenoviral CRISPR/Cas9 genome editing tools can also be used to deliver Cas9 and DNA templates in ... CRISPR/Cas9 research! To find more information about the adenoviral delivery of CRISPRS/Cas9 using the... also Adeno Cas9 and Adeno EA. Or if you're looking for a broader range of CRISPRs plasmid tools, find...Genome Editing of Pten by CRISPR/Cas9 in Mouse Liver in Spite of Cas9-Specific Immune Responses. Human...

  13. CRISPR 101: Epigenetics and Editing the Epigenome

    Type
    Blog Post
    ...Staphylococcus aureus Cas9 (Sa-Cas) and deposited plasmid IF311: pMAGIC (R4-R3) NLS-Sa dCas9-NLS-LSD1. LSD1 ...types. They have also fused LSD1 to x-Cas9(3.7). x-Cas9(3.7) is a SpCas9 mutant that exhibits increased PAM... Find plasmids from this publication at Addgene. Additional Resources on the Addgene Blog Cas9 Activators...pCMV-dCas9-D3A) and Tet-dependent (TetO-dCas9-D3A) constructs. For lentiviral expression, Fuw-dCas9-Dnmt3a...deposited mammalian expression constructs including pcDNA-dCas9-p300 Core and pcDNA3.3-Nm-dCas9-p300 Core, as...as well as pLV-dCas9-p300-P2A-PuroR for lentiviral expression. Figure 1: dCas9-p300 adds H3K27ac... created pdCas9-Tet1-CD for targeted cytosine demethylation in mammalian cells. This plasmid is used with...

  14. CRISPR 101: Targeting RNA with Cas13a (C2c2)

    Type
    Blog Post
    ...Abudayyeh et al., 2016). In the endogenous CRISPR/Cas9 system, Cas9 uses both a tracrRNA and crRNA to facilitate...multiplexing; expressing 5 gRNAs simultaneously produces comparable gene knockdown to single gRNA expression. Although... 27, 2020. CRISPR, and specifically Cas9 from S. pyogenes (SpCas9), is truly an exceptional genome engineering...Perhaps the biggest difference between Cas13a and Cas9 is that Cas13a binds and cleaves RNA rather than...enzymes - Cas13a contains two HEPN domains, whereas Cas9 uses HNH and RuvC domains to cleave target DNA. ...roles for HEPN domains in other proteins. As with Cas9, mutating key residues in the Cas13a molecule results... Guide RNA Target PAM (PFS) Cleavage Mechanism Cas9 HNH, RuvC TracrRNA, crRNA DNA 5' NGG Specific...

  15. Hot Plasmids - March 2019 - Anti-CRISPR, 2in1 Cloning, Fluorescent Voltage Indicators, and Photoswitchable Proteins

    Type
    Blog Post
    ... of protein coexpression. By introducing a single plasmid containing both GOIs, expression levels are ...highlight a subset of the new plasmids in the repository through our hot plasmids articles. These articles ...recent plasmid deposits and we hope they'll make it easier for you to find and use the plasmids you need... recent plasmid deposit please sign up here.          Listen to this episode of Hot Plasmids! Optogenetic...genes-of-interest (GOI) into two independent expression cassettes on a single plasmid (Grefen and Blatt, 2012). Similar...DNA transfer, each GOI is usually expressed from a different plasmid, which may lead to unequal gene dosage...Grefen and Blatt, 2012). The 2in1 Plasmid Toolkit consists of 4 plasmid toolkits generated with 4 different...

  16. Hot Plasmids - June 2019 - Optogenetics, Acoustic Reporter Genes, microRNAs, and the CRISPR-Cas9 system CHIME

    Type
    Blog Post
    ... (ARG) plasmids to better monitor gene expression in vivo using ultrasound. These ARG plasmids contain...pXPR_053. They expressed this construct in bone marrow stem cells from Cas9-expressing mice, and used ...highlight a subset of the new plasmids in the repository through our hot plasmids articles. These articles ...recent plasmid deposits and we hope they'll make it easier for you to find and use the plasmids you need... recent plasmid deposit please sign up here.      Listen to this episode of the Hot Plasmids podcast ...responses in C. elegans. Find the split BoTN/B plasmids at Addgene. Liu Q et al., Neuron. 2019. PubMed... microbe travels through a host organism, the expression of some genes can change depending on the location...

  17. Hot Plasmids and Viral Preps - March 2021

    Type
    Blog Post
    ...subset of the new plasmids and viral preps in the repository through our hot plasmids articles. These articles...recent plasmid deposits and we hope they'll make it easier for you to find and use the plasmids you need...-baToN, “sender” cells express GFP on the surface and “receiver” cells express an anti-GFP nanobody on...response 1 (TIR1) expressed under a desired promoter with a specific spatiotemporal expression pattern and ...highlights from recent CRISPR plasmids. To find all of the CRISPR plasmids available from Addgene, head...head over to our CRISPR Plasmids and Resources page.  Plasmids for delivering CRISPRi components by a single.... Once activated, Cas9 cleavage can induce DSBs within minutes. New CRISPR plasmids for Borrelia burgdorferi...

  18. Fluorescent Tagging of Endogenous Genes with SapTrap

    Type
    Blog Post
    ...co-injection of the targeting vector and a Cas9 expression plasmid will insert the desired genetic tag and...as a universal donor plasmid. The CRISPaint toolkit includes several donor plasmids for protein tagging...are then mixed and transfected along with a Cas9 expression vector. They can also be introduced directly...directly into a Drosophila S2 cell line stably expressing Cas9. This PCR toolkit offers C- and N-terminal tagging...this system lies in the ability of the endonuclease Cas9 to create a DNA double stranded break (DSB) at a...The Jorgensen Lab recently developed a modular plasmid assembly toolkit called SapTrap to improve CRISPR...accepts the sgRNA target sequence for U6 promoter expression and the second site accepts the homology arm ...

  19. Genome-wide Screening Using CRISPR

    Type
    Blog Post
    ...in a 1-plasmid system, in which Cas9 is included on the gRNA-containing plasmid, or a 2-plasmid system...CRISPR/Cas9 over previous genome editing techniques is its simplicity and versatility. CRISPR/Cas9 consists...gRNA-containing plasmids, each with a different genomic target. The goal of a CRISPR screen is to use Cas9 and a...to deliver a pooled population of gRNAs to Cas9 expressing cells. Pooled lentiviral CRISPR libraries (...(A) to be used to generate lentivirus (B). Cas9-expressing cells or wild-type cells are treated with lentivirus...contain thousands of gRNA plasmids, discerned only by a unique barcode on each plasmid. As such, sequencing... how CRISPR/Cas9 can be used in other types of experiments, check out our CRISPR Plasmids and Resources...

  20. New Tools Enable CRISPRa for Neuroscience Applications

    Type
    Blog Post
    ...ubiquitous expression of sgRNA(s) and mCherry (vector 1) with neuron-specific expression of dCas9-VPR (Figure...system with ubiquitous expression of sgRNA (top) and neuron-specific expression of dCas9-VPR (bottom). Created...et al., 2018). This mouse expresses a modified version of the “Suntag” Cas9 activator that recruits up... for generating the SPH mouse. (B) AAV plasmids used to express CRE under neuron-specific promoters (aCaMKII... (i.e., how many sgRNA expression cassettes will fit). Cre-dependent expression of the SPH activator paired...neuronal populations.  The Yang Lab’s AAV plasmids used to express Cre under neuron-specific promoters, including...Figure 2), a popular Cas9 activator developed by the Church Lab. Neuron-specific expression is mediated by ...
Showing: 21 - 40 of 224 results