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We narrowed to 150 results for: cas9 expression vector

Showing: 101 - 120 of 150 results

  1. Tips for arabidopsis transformation

    Type
    Blog Post
    ...Addgene  Choosing an expression system Choosing an appropriate expression vector is the first step in .... (2007) ‘Improved Gateway Binary Vectors: High-Performance Vectors for Creation of Fusion Constructs ...from the Tsuyoshi Nakagawa lab are binary gateway vectors and can be used for promoter swapping and generating...for visual selection of transformants based on expression of RFP in the seed (Emami, Yee and Dinneny, 2013...many insertions can cause artifacts from over-expression and it is likely that such a construct will be... ‘A robust family of Golden Gate Agrobacterium vectors for plant synthetic biology’, Frontiers in Plant...sweet potato contains Agrobacterium T-DNAs with expressed genes: An example of a naturally transgenic food...

  2. CRISPR Plasmids - Tagging

    Type
    Collection
    ...Plasmids: pX330S-2-PITCh - expresses Cas9 & PITCh-gRNA pX330A-FBL/PITCh - expresses Cas9, PITCh-gRNA, & FBL-specific...targeting vector assembly, and the remaining 5 plasmids include Cre and FLP expression vectors, a general... one vector from the Doyon lab, eSpCas9(1.1)_No_FLAG_AAVS1_T2 (Addgene #79888) , which expresses an untagged...plasmid-system consists of two components: A vector containing Cas9 and a validated gRNA, based on the Zhang...consists of two components: An all-in-one CRISPR/Cas9 vector based on the Zhang lab's PX330 , simultaneously...gene-specific CRISPR and donor vectors as well as pX330S-2-PITCh, expressing a generic gRNA targeting the... that uses PCR to generate both an expression cassette for the Cas9-programming sgRNA and HR donors for...

  3. CRISPR Plasmids - Prime Edit

    Type
    Collection
    ...contains Cas9. ID Plasmid Expression System Promoter Guide RNA Type Cloning Enzyme Co-expressed Cas9 Selection... top Empty Prime Editing gRNA Vectors A selection of empty gRNA vectors suitable for prime editing are...below. Use the search bar to find a gRNA vector based on expression system, promoter, the type of gRNA (e.g...transcriptase (RT) is fused to the C terminus of Cas9 H840A nickase. The fusion enzyme is capable of installing...template. During prime editing, the pegRNA directs the Cas9 nickase to the target sequence where it nicks the...nuclear localization signals, and two mutations in Cas9 to enhance nuclease activity; can be used in PE2...PE6 — small prime editors with optimized RT and/or Cas9 domains PE7 — addition of RNA-binding exonuclease...

  4. Cloning Mammalian Cells with the Agarose Method

    Type
    Blog Post
    ...Fluorescent Protein Plasmids Find Mammalian Expression Vectors ...that overexpress genes of interest; more recently we have also been using Addgene CRISPR vectors to generate... enzyme assays to select both over- and under-expressing clones. If GFP-encoding cDNA is co-transfected...well plate for further analysis. To assess CRISPR/Cas9 knockout efficacy, we have used both Western blotting...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...

  5. Hot Plasmids: Fall 2025

    Type
    Blog Post
    ...cells express iCre with a mitochondrial anchor and a TEV cleavage site. Recipient cells express DsRed ...killswitch (GFP-nb–KS) is delivered using a vector that also expresses mCherry to mark transfected cells. The...Affordable, low-tech CRISPR/Cas9 experiments By Ashley Waldron CRISPR/Cas9 has revolutionized biotechnology...chromoprotein and gRNA plasmids. Nuclease-dead Cas9 (dCas9) protein is supplied separately. Results can ...cell-surface proteomes (Wu et al., 2025). The team expressed extracellularly-anchored horseradish peroxidase...nuclease, and excises the DsRed-STOP to trigger expression of EGFP. Figure 2: Schematic of the MitoTRACER...Confocal image of CA2 neurons in a mouse brain slice expressing PinkyCaMP; scale bar 50 µm. Reproduced from Fink...

  6. AAV Q&A with Tim Miles

    Type
    Blog Post
    ...This will be highly dependent on vector dose. The color diversity in Vector Assisted Spectral Tracing (VAST...drop in transduction coverage compared to just one vector.   Q2: Do you think it is possible to make an efficient...this scenario, you may be best served by a dual vector, split intein approach.   Q3: What’s the best method...for model organisms.    Q11: How consistent is expression is when injecting CAP-B10 or CAP-B22 in marmosets...primer for beginners Adenoviral delivery of CRISPR/Cas9 Tips for a first time AAV user Resources on Addgene.org...Plasmids Adenovirus Guide Page Subscribe to Viral Vectors Blog Posts ...

  7. CRISPR Guide

    Type
    Collection
    ...more about Cas9 multiplexing . Browse Plasmids: Multiplex gRNA Vectors Engineering the Cas9 Endonuclease... available with multiple Cas9 variants, including high fidelity Cas9s, Cas9s with various PAM requirements...lentiviral transfer vectors (Figure 8B). Libraries come in 1-vector systems, in which Cas9 is included in ...a 2-vector system, you’ll need to either co-infect with a Cas9-producing lentivirus or use a Cas9-expressing...strand SpCas9-HF1 — disrupt Cas9’s interactions with DNA phosphate backbone HypaCas9 — increase Cas9 proofreading...more Cas9 fusion tools later in the guide. Figure 3: Overview of Cas9n. Increasing fidelity of Cas9 Researchers...enhanced specificity by engineering Cas9 into high fidelity Cas9s (hfCas9) . There is currently no defined...

  8. Custom CRISPR Screens & the Green Listed Software

    Type
    Blog Post
    ...screens Differentially expressed genes  As the costs for global gene expression analyses like RNAseq are...test which of the differentially expressed genes found in a gene expression dataset are causal for a phenotype...differentially expressed genes that are known drug targets, or all differentially expressed genes that code...sequencing to identify which gRNAs are differentially expressed in the separate groups and, as a consequence, ...CRISPR screens designed to target differentially expressed genes can be the next stage in making sense of...interactions to a large extent involve receptors expressed on the cell surface. Thus, a screen can be used...The gRNA oligos are subsequently cloned into your vector of choice using the adapter sequences that you ...

  9. Overcoming the Challenges of Lentiviral Production

    Type
    Blog Post
    ... While lentiviral vectors are popular gene delivery tools, producing lentivirus, can pose certain challenges... a usable titer, or fine-tuning selection and expression in your target cell line, researchers often find...transcriptional activation system to drive high levels of expression of certain viral components; according the manufacturer...single 72h harvest is best for larger genes such as Cas9. See figure 1 for an example of how changing transfection...determination of the packaging limit of lentiviral vectors. Hum Gene Ther. 2001 Oct 10;12(15):1893-905. PubMed...Tips for titering your lentivirus Visit the viral vector featured topic page Resources on Addgene.org ...Guide Find lentivirus plasmids Browse all viral vectors ...

  10. Treating Muscular Dystrophy with CRISPR Gene Editing

    Type
    Blog Post
    ...long period of time. High fidelity Cas9s such as eSpCas9 and SpCas9-HF should be explored to reduce off-target...myoblasts; when injected into DMD mice, these cells expressed functional dystrophin. Editing genes in vivo is..., the CRISPR components would only need to be expressed for a short period of time, which would prevent...et al. show that the percentage of dystrophin-expressing muscle cells increases over time, and all three...three groups confirm dystrophin expression via Western blot, at <10% of the wild-type level. Muscle histology...stem cells 2. Remove any immunogenicity of the AAV vector Once delivery has been optimized, it’s important...off-target editing. Short-term CRISPR expression would be especially desirable, as it would reduce the...

  11. Click Biology: How to Click and Play to Explore Biology

    Type
    Blog Post
    ...fluorescent ligands to HaloTag PCV-Cas9, Cas9-PCV HUH-tag Cas9 plasmids with N- or C-terminal PCV2 HUH-tag...transactivator to increase fluorescent protein expression C234-E25: attB2r-SNAP-attB3 SNAP-tag Gateway...can also assemble vaccines, target gene therapy vectors, or lock together smart biomaterials. There are...

  12. Plasmids 101: Inducible Promoters

    Type
    Blog Post
    ...inducible system is found in many bacterial expression vectors. Negative inducible promoter pBad is another...gene, it therefore determines the timing of gene expression and largely defines the amount of recombinant...Temperature inducible promoters Temperature sensitive expression systems are typically less leaky than chemically...chemically induced promoters; they show near-zero expression at regular temperatures but can be induced by ... promoters, in which a gene of choice is only expressed following exposure to a brief heat shock. In the...depositors have developed heat shock-inducible Cre and Cas9 for easy genome engineering in species like C. elegans...promoters Light is another way to activate gene expression, and two-component systems used in synthetic ...

  13. AAV Molecular Tools

    Type
    Collection
    ...encode Cas9 nuclease used in the CRISPR-Cas9 system for targeted genome editing. ID Name Expression System...AAV-CMV::NLS-SaCas9-NLS-3xHA-bGHpA CMV-driven, constitutive Expression of human codon-optimized Cas9 from Staphylococcus... Viral Vector Packaging Service AAV Molecular Tools Viral Vector Packaging Service: Molecular...with tetracycline (tet)-inducible expression systems . ID Name Expression System Activity Serotype PI 99118...transsynaptic tracer protein to express in presynaptic glutamatergic neurons that express Cre, Flp payload. Must...Cre-dependent expression of ER-localized TurboID (with HA and V5 tag). 8 Jonathan Long Overexpression Tools These...These AAV encode various genes for overexpression. ID Name Expression System Activity Serotype PI 58376...

  14. Hot Plasmids and Viral Preps - September 2021

    Type
    Blog Post
    ...of robust expression of high G+C content genes, as validated through monitoring the expression of fluorescent...different vectors and flow cytometry is used to sort populations by mCherry and GFP expression to check...end-point sequencing assays. These plasmids use “Cas9-mediated adenosine transient reporter for editing... be seen through the success of the change in expression of GFP, for the stop codon should have changed...

  15. Fluorescence Titering Assay

    Type
    Protocol
    ...pHAGE-TO-dCas9-3XmCherry . 72 h post transduction, cells were assayed for mCherry expression using fluorescence...Assay for Lentivirus You may also like... Viral Vector Guides Virus Blog Posts Mol Bio Protocols Viral...marker. Determining the titer of your lentiviral vector allows you to control the multiplicity of infection...protocol was developed using low titer lentiviral vectors (10 5 TU/mL). If you anticipate that your viral...

  16. CRISPR Plasmids - dCas9-FokI

    Type
    Collection
    ...Purify Tag Visualize dCas9-FokI Screen Pooled Libraries gRNAs Empty gRNA Vectors By Species Mammalian ...Tools CRISPR Blog Posts A catalytically inactive Cas9 (dCas9) is fused to FokI nuclease. When FokI dimerizes...CRISPR-FokI plasmids. Plasmids are available for expression in mammalian systems and Drosophila. Mammalian... CRISPR dCas9-FokI CRISPR: dCas9-FokI...with dCas9 fused to FokI for creating double-strand breaks (DSBs). CRISPR...gRNAs, binding ~15-25 bp apart, are required for dCas9-FokI to dimerize at a given region of the genome...

  17. How to Deposit Your Plasmids with Addgene

    Type
    Blog Post
    ...directing CRISPR/Cas9 to a targeted genomic location, while shRNAs are used to knockdown expression of mRNAs ...the vector backbone you cloned your insert into and fill out the appropriate boxes with the vector backbone...backbone manufacturer, vector backbone size, total size of your vector + insert, any modifications you ...insert in E. coli, then this is a Bacterial Expression vector. If your plasmid can be classified in multiple...lentiviral vector to be used in mammalian cells, check Lentiviral and Mammalian Expression). This information...provide any full or partial plasmid sequences and any vector maps. The more sequence data available, the better... was a part of the insert (when cloned into the vector backbone). If your plasmid has more than one insert...

  18. Hot Plasmids - November 2023

    Type
    Blog Post
    ...fluorescently labeled. Using lentiviral vectors to stably or inducibly express tagged iPAK4 from different promoters...edits, such as long or highly structured templates. Cas9 variants PE6e–g often had even higher editing efficiencies...to reprogram delivery vehicles like lentiviral vectors, CreVLPs, and eVLPs for in vivo and in vitro applications...activation events as bands of induced GFP-iPAK4 expressed under promoters like the tet-responsive element...to create a three-dimensional atlas of protein expression in the adult mouse brain using thick-section ...

  19. CRISPR 101: Off-Target Effects

    Type
    Blog Post
    ...blog CRISPR 101: Which Cas9 do I choose for my CRISPR experiment? CRISPR 101: Cas9 The Other Cas(s) CRISPR...cleave DNA and instead activates/represses gene expression by various mechanisms. The RNA guide is typically...- GUIDE-Seq and integrase-defective lentiviral vectors - which rely on insertion of DNA by the NHEJ pathway...Ma X, Gao F, Guo Y. Off-target effects in CRISPR/Cas9 gene editing. Front Bioeng Biotechnol. 2023 Mar ...specific variants. HypaCas9, sSpCas9(1.1), SpCas9HF1, and evoCas9 are a few spCas9s which have been evolved...of Maya Kostman. The most commonly used Cas, spCas9, has gone through multiple rounds of optimization...

  20. PRIDICT: Predicting Efficiencies of Prime Editing Guide RNAs

    Type
    Blog Post
    ...Edit Collection The CRISPR Guide Empty gRNA Expression Vectors...2023). This method uses a prime editor (typically a Cas9 H840A nickase fused to a reverse transcriptase) ...Steensel, B. (2021). Impact of chromatin context on Cas9-induced DNA double-strand break repair pathway balance...Yoon, S., & Kim, H. H. (2019). SpCas9 activity prediction by DeepSpCas9, a deep learning–based model with... and PE5 prime editing strategies) based on a DeepSpCas9 score ranking (Kim et al., 2019). PRIDICT saves...
Showing: 101 - 120 of 150 results