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We narrowed to 117 results for: cas9 expression vector

Showing: 81 - 100 of 117 results

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

    Type
    Blog Post
    Published
    Sept. 28, 2020, noon
    ...technology (Ronda et al., 2016). pMA7CR_2.0 expresses lambda Red and Cas9, which are separately inducible by L-arabinose...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...targeting either a catalytically dead Cas9 (dCas9) alone or a dCas9 fused to the omega subunit of RNA polymerase...tuberculosis. dCas9 and a gRNA are expressed on pRH2502 and pRH2521, respectively. Both dCas9 and the gRNA... from Addgene consists of two plasmids: pCas9: carries Cas9 and chloramphenicol resistance pCRISPR: carries...

  2. Hot Plasmids - October 2020

    Type
    Blog Post
    Published
    Oct. 13, 2020, 1:15 p.m.
    ...from other regions of the cell. By combining vectors expressing different fluorescent proteins targeting ...Applications (CHyMErA) Knockout Libraries express hybrid Cas9-Cas12a gRNAs under a single U6 promoter. ...display (FingRs), Xue Han’s lab has developed AAV vectors to selectively visualize excitatory and inhibitory...excitatory and inhibitory synapses, respectively. Once expressed in the neuron, the FingRs bind to the endogenous.... The team engineered C. elegans cell lines to express wrmScarlet1-10 unattached to any other cellular...proliferation in acute myeloid leukemia and saw that expression of H2B-FT consistently predicted the cell cycle...different cell types. In one example, H2B-FT was expressed in murine B cells (BaF3), which depend on IL-3...

  3. Generating Mouse Models Using CRISPR/Cas9

    Type
    Blog Post
    Published
    July 12, 2016, 2:30 p.m.
    ...alternative Cas9, from Streptococcus aureus, is as effective at editing mouse embryos as Cas9 from S. pyogenes...., 2016). AAV can also be used as a vector to deliver CRISPR/Cas9 either through injection into either... Knockin by CRISPR/Cas9-Mediated Genome Editing with Adeno-Associated Viral Vector. iScience 9:286–297...guest bloggers, Wenning Qin and Haoyi Wang. CRISPR/Cas9 is revolutionizing the mouse gene-targeting field...ever to manipulate the mouse genome using CRISPR/Cas9. Similar to the human genome, the mouse genome ...robust in performance. In its natural setting, CRISPR-Cas9 is an acquired immune system in bacteria and archaea...) of about 125 nt that specifies the target, the Cas9 endonuclease that creates the DNA double-strand ...

  4. How to Design Your gRNA for CRISPR Genome Editing

    Type
    Blog Post
    Published
    Sept. 24, 2020, 1:15 p.m.
    ...is_json="true" %}{% raw %}"Browse Empty gRNA Expression Vectors"{% endraw %}{% end_module_attribute %}{% ... capacity of a locus. When using S. pyogenes Cas9 (SpCas9), potential target sites are both [5’-20nt-NGG... (inhibition) technologies – a nuclease-dead Cas9 (dCas9) is directed near the promoter of a target gene...href":"https://www.addgene.org/crispr/empty-grna-vectors/","type":"EXTERNAL"},"open_in_new_tab":true,"no_follow..., a specific base edit, or modulation of gene expression. Location and sequence are important considerations...with CRISPR technology is usually accomplished by Cas9-mediated dsDNA breaks: following a cut, the error-prone...even more exquisite for the so-called base editor Cas9, which makes DNA changes in the absence of dsDNA...

  5. Four Ways to Package Transgenes That Exceed the Size Limit of Adeno-associated Virus

    Type
    Blog Post
    Published
    Sept. 1, 2020, 1:15 p.m.
    ...ideal split vector system. Overall the efficiency of transgene expression from split vectors has been inconsistent... overviews of three split vector strategies plus one additional split vector strategy for increasing the...homology directed repairs (HDR). Split AAV vector approaches Split vector approaches increase the size of the...Generally an overlapping split vector has equal or higher transgene expression when compared to using a single...copies of both vectors. Examples of use: Halbert et al., 2002 used an overlapping vector with 440 or 1000bp...dystrophy. While mini-dystrophin expression levels were lower than expression of wild-type dystrophin in normal...to deliver LacZ to muscle in mice. Expression from the hybrid vector, as measured by β-galactosidase activity...

  6. Reaching out to China: Canton Nucleic Acids Forum (CNAF) 2015

    Type
    Blog Post
    Published
    Dec. 1, 2015, 3:30 p.m.
    ...using bacterial phage vectors with CRISPR/cas9 sensitizing gene cassettes. Thse vectors are engineered to ... tetracycline cas9 inducible knock-out system. His lab is also exploring the use of cas9 proteins from... closed RNAs seem to be conserved in mammals, expressed in neurons, and are especially abundant in neuronal...There are many clever ways labs are using CRISPR/cas9 and more are being reported. Zhou Songyang from ...

  7. Quick Guide to All Things Lentivirus

    Type
    Blog Post
    Published
    March 21, 2017, 2:30 p.m.
    ...(tTA, also expressed by your transfer vector) binds to the TRE and activates the expression of your gene...your experiment using the lentiviral vector system. Viral vectors have been increasingly popular in fundamental...primary cells. Amongst the different vectors used, lentiviral vector constructs have proven very useful ...properties make lentiviral vectors fantastic options for delivering shRNA, CRISPR/Cas9 components, and fluorescent...fluorescent sensors. Retroviral vector background Until the development of viral vectors, gene transfer into mammalian...un-transfectable cells, labs use viral vectors and particularly retroviral vectors because they target a broader...attractive choice for a gene transfer vector. Lentiviral vectors Although retroviruses are still in use...

  8. Addgene-Seeding Labs Plasmid Grant: Accelerating Science Globally - 5 Scientists, 4 Countries & 3 Continents!

    Type
    Blog Post
    Published
    July 18, 2017, 12:41 p.m.
    ...the light‐inducible bacterial expression system pDawn to clone and express newly identified proteins and...project the expression of isolated bacterial proteins can be achieved using a bacterial expression system ...dietary supplements! Development of a bacterial expression system for renewable biotechnology products Dr...plant, Dr. Robertson aims to make use of the CRISPR/Cas9 technology to engineer plants that are able to destroy...focuses on molecular biotechnology for improving expression of nutritionally and therapeutically significant...cannot produce them. By using yeast integration vectors (see for example this yeast cloning kit) she intends...system such as the pET GFP LIC cloning vector and the pOKD plasmids. The recombinant Cry proteins will then...

  9. Hot Plasmids: Summer 2025

    Type
    Blog Post
    Published
    Aug. 28, 2025, 1:15 p.m.
    ...be carried by viral vectors (~400 amino acids, less than a third of the size of Cas9). Figure... mGold2. Top: TIRF-SIM imaging of COS-7 cells expressing LifeAct fused to mVenus, mGold2s, or mGold2t....Figure 2: Comparison of TnpB (A) and Cas9 (B) genome editors. Effector nucleases are shown bound to the target..., recruited by either the ωRNA (TnpB) or a gRNA (Cas9). Drawings not to scale. Created with BioRender....thousands of eligible AAV plasmids to order viral vectors ready to use in your lab. Find Addgene’s AAV6 and... synapses. In addition, ATLAS itself could be expressed Cre-dependently to allow tracing exclusively from...

  10. Adenoviral Vector Production and Troubleshooting

    Type
    Blog Post
    Published
    June 14, 2018, 11:44 a.m.
    ...Adenoviral vectors (AdV) are attractive vectors for research applications and gene therapy: they can ...Three generations of adenoviral vectors: the first generation vectors are deficient in the E1 and E3 regions... second generation vectors are deficient in E1, E2, E3, and E4. Gutless Ad vectors (helper-dependent (...creating adenoviral vector constructs. It consists of 2 plasmids: a shuttle vector (in which the transgene...death of transduced cells and loss of transgene expression (Interestingly, the strong immunogenicity of ...Adenovirus type 5. Three generations of adenoviral vectors The adenovirus virion is a non-enveloped particle...capacity is limited to 8.2 kb, and in vivo transgene expression ceases relatively quickly due to immune response...

  11. Tips for arabidopsis transformation

    Type
    Blog Post
    Published
    Oct. 25, 2018, 1:23 p.m.
    ...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...

  12. Cloning Mammalian Cells with the Agarose Method

    Type
    Blog Post
    Published
    Sept. 7, 2017, 12:17 p.m.
    ...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...

  13. Hot Plasmids: Fall 2025

    Type
    Blog Post
    Published
    Nov. 11, 2025, 2:14 p.m.
    ...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...

  14. AAV Q&A with Tim Miles

    Type
    Blog Post
    Published
    May 17, 2022, 1:15 p.m.
    ...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 ...

  15. Custom CRISPR Screens & the Green Listed Software

    Type
    Blog Post
    Published
    July 11, 2017, 2:30 p.m.
    ...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 ...

  16. Overcoming the Challenges of Lentiviral Production

    Type
    Blog Post
    Published
    Aug. 7, 2018, 12:31 p.m.
    ... 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 ...

  17. Treating Muscular Dystrophy with CRISPR Gene Editing

    Type
    Blog Post
    Published
    Jan. 26, 2016, 3:30 p.m.
    ...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...

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

    Type
    Blog Post
    Published
    Oct. 21, 2025, 1:15 p.m.
    ...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...

  19. Plasmids 101: Inducible Promoters

    Type
    Blog Post
    Published
    Jan. 18, 2018, 2:34 p.m.
    ...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 ...

  20. Hot Plasmids and Viral Preps - September 2021

    Type
    Blog Post
    Published
    Sept. 21, 2021, 1:15 p.m.
    ...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...
Showing: 81 - 100 of 117 results