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We narrowed to 117 results for: cas9 expression vector

Showing: 101 - 117 of 117 results

  1. How to Deposit Your Plasmids with Addgene

    Type
    Blog Post
    Published
    Feb. 27, 2024, 2:15 p.m.
    ...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...

  2. Hot Plasmids - November 2023

    Type
    Blog Post
    Published
    Nov. 14, 2023, 3:35 a.m.
    ...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 ...

  3. CRISPR 101: Off-Target Effects

    Type
    Blog Post
    Published
    Feb. 8, 2024, 2:15 p.m.
    ...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...

  4. PRIDICT: Predicting Efficiencies of Prime Editing Guide RNAs

    Type
    Blog Post
    Published
    June 25, 2024, 1:15 p.m.
    ...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...

  5. RNA Interference in Plant Biology: New Tools for an Old Favorite

    Type
    Blog Post
    Published
    Oct. 27, 2020, 1:15 p.m.
    ...transgene. Typically, the desired target of RNAi is expressed as inverted repeats separated by a loop region...considerations, such as the starting material (DNA-based vector or purified RNA) and the initial state of the dsRNA... easiest approach to induce RNAi involves the expression of long hairpin RNA (hpRNA). This method is facilitated...construct has unique features, such as inducible expression of the hpRNA and/or fusing your gene target sequence... Although downstream work is required, several vectors can be found at Addgene that could serve as an ...an excellent start (#22988, #137884). Both vectors allow the creation of amiRNAs based upon miRNA precursors...Bassik MC (2016) Systematic comparison of CRISPR/Cas9 and RNAi screens for essential genes. Nat Biotechnol...

  6. What's Your Organism? Expanding Genomic Tools via the NSF EDGE Program

    Type
    Blog Post
    Published
    May 2, 2019, 12:41 p.m.
    ...   It’s not an overstatement to say that CRISPR/Cas9 editing is one of the key tools now providing access...continues to be challenging. Organism specific viral vector systems would also be beneficial for many systems...snail can be manipulated by injected RNA and can express fluorescent proteins. This lab has created an automated...-Mediated Ovary Transduction of Cargo)  delivers Cas9 RNP to the arthropod germline by injection into ...

  7. Evolution of Brainbow: Using Cre-lox for Multicolor Labeling of Neurons

    Type
    Blog Post
    Published
    April 24, 2015, 2:39 p.m.
    ...CRISPR-Cas9 genome editing may be the hot new way to manipulate gene expression, but other gene manipulation...recombination, RFP is expressed. Cre can mediate one of two deletions to allow YFP or M-CFP expression; these deletions...ways to spatially and temporally control gene expression, especially in in vivo models, and new Cre-lox...fluorescent protein in the construct, OFP (orange), is expressed when Cre is absent; stochastic Cre excision results... and inversion. The Brainbow-2.1 construct can express one of four colors (n-GFP, RFP, YFP or M-CFP.) ...remaining cassette can invert as long as Cre remains expressed; once Cre is removed from the system, the fluorescent...coding sequence is closest to the promoter will be expressed. This system differs from Brainbow-1.0 in that...

  8. Viral Vectors 101: Parts of the AAV Transfer Plasmid

    Type
    Blog Post
    Published
    Aug. 11, 2020, 1:15 p.m.
    ... of expression. For example, CAG is a strong promoter that drives very high, ubiquitous expression, while...such as SaCas9, CRISPR gRNAs, or your gene of interest. Promoter The promoter drives the expression of the...outside of the ITRs does not get packaged in the AAV vector. The cloning capacity between the ITRs is ~4.4 ...type-specific, which can help you restrict the expression of your transgene to a specific species or location...to spatially and temporally control transgene expression. Lox sites are directional 34 bp sequences that...nuclear export of mRNA and helps increase the expression of your transgene. The WPRE is located at the...design a construct for the successful and specific expression of your gene of interest. References Haery ...

  9. Hot Plasmids - August 2020

    Type
    Blog Post
    Published
    Aug. 18, 2020, 1:15 p.m.
    ...approaches rely on continual expression of both dCas9 and the gRNA. This expression can vary significantly ...piggyBac vectors for uniform and sustained expression of single or multiplexed gRNAs and of dCas9 fusions...and track cells expressing both dCas9 and gRNAs. The multi-gRNA piggyBac vector integrates into ...easier to clone, express, and purify compared to antibodies. Nanobodies are easily expressed in the cytosol...#140702) The pMAK461-64 plasmids were expressed in SHuffle T7 Express competent E.coli (NEB) using IPTG induction... contain an EGFP reporter. Expression of the dCas9-KRAB activator or dCas9-VPR repressor in hPSCs is also... the mCitrine gene. mCherry is constitutively expressed. Cells are then cultured and FACS sorted. Image...

  10. The Fluorescent Vegetables in Aptamer Soup

    Type
    Blog Post
    Published
    Jan. 29, 2019, 2:27 p.m.
    ...SELEX, the RNA pool was cloned into bacterial expression vectors to create a library. Bacteria were transformed...leading to conformational changes that control the expression of coding RNA. Spinach-based riboswitches can...cellular RNA to specific intracellular locations using Cas9 and track the localization of the complex. Broccoli...quantify RNA transcription by RNA Polymerase III. By expressing Corn under promoters for transcripts uniquely...

  11. Split Fluorescent Proteins for Studying Protein-Protein Interactions

    Type
    Blog Post
    Published
    May 3, 2022, 1:15 p.m.
    ... vitro, and can be used for measuring protein expression or localization in cells, or even single-particle...Mol Syst Biol. 2014 Galit Lahav EYFP Gateway vectors with EYFP split at 175 Arabidopsis homolog of the...Split-horseradish peroxidase (Alice Ting Lab 2016)  Split-Cas9 for inducible genome editing (Feng Zhang Lab 2015...

  12. Antibody Validation for Flow Cytometry

    Type
    Blog Post
    Published
    Sept. 10, 2024, 1:15 p.m.
    ...of interest when expressed at endogenous levels (Laflamme et al., 2019). CRISPR-Cas9 approaches are commonly...flow cytometry, and mRNA expression.   If your protein of interest is expressed at similar levels in ...useful to produce a sample with mixed expression, with the expression measurable through an epitope tag such...epitopes. Detection of overexpressed (tagged or untagged) protein When expression plasmids are available...differences between endogenous expression and transfected overexpression, this approach cannot confirm ...caused by overexpression. Additionally, if the cell line used has endogenous levels of expression, this can...preferred. The overexpression approach is therefore best suited to cell lines without any expression of the target...

  13. Fluorescent Proteins 101: Monitoring Cell Mobility Using Fluorescent Proteins

    Type
    Blog Post
    Published
    Aug. 15, 2017, 1:24 p.m.
    ...loxP sites such that their expression is turned on or off in cells expressing Cre. The result of a system...allows the visualization of fluorescent proteins expressed in the specimen.  2. Fluorescence Activated Cell...immune lineage, scientists could easily track GFP expressing cells. For example, by harvesting lymph nodes...efforts have been made to create transgenic mice expressing fluorescent protein reporter constructs in the...above depicts the infiltration of neutrophils expressing LysM-GFP (green) into a draining lymph node after...other technologies such as optogenetics and CRISPR/Cas9 will help immunologists create even better tools...Subcellular Localization Find Fluorescent Protein Empty Vectors ...

  14. Plasmids 101: Screening Strategies Used in Plasmid Cloning

    Type
    Blog Post
    Published
    Sept. 15, 2020, 1:15 p.m.
    ...bacteria that have taken up this vector, but it won’t tell you whether the vector contains the gRNA insert or...X-gal allows you to identify cells that took up the vector and distinguish between plasmids that contain the...𝛽-galactosidase. Complementing this mutation by expressing this deleted portion, for example on a plasmid...functional 𝛽-galactosidase. Blue-white screens rely on vectors where the gene of interest is inserted into the..., you will see two bands: one representing your vector backbone and the other representing the insert....doesn’t contain the insert, you will just see the vector backbone band. Or, you can choose an enzyme that...specific to your insert or by using primers that are vector specific but amplify a potential insert. If you...

  15. Hot Plasmids: Summer 2024

    Type
    Blog Post
    Published
    Aug. 27, 2024, 1:15 p.m.
    ...much smaller proteins than SpyCas9, allowing them to be packaged in AAV vectors, potentially with multiplexed...al., 2024). In MagIC-cryo-EM, target proteins (expressed as a GFP fusion in this proof-of-concept study...showed that AAV-delivered CHARM could silence the expression of prion protein in mouse brains for at least...gene product; it simply turns the target protein expression off. Plus, DNA methylation is a long-lasting ...-silencing CHARM variants that stop their own expression after silencing their target — potentially improving...by targeting marker proteins that are uniquely expressed in a given cell type. For example, glial fibrillary...GFAP) is a glia-specific intermediate filament expressed in astrocytes throughout the brain, making it ...

  16. Interview: Hodaka Fujii on enChIP, New CRISPR Tools, and More

    Type
    Blog Post
    Published
    Dec. 2, 2014, 7:23 p.m.
    ...We generated a series of retroviral vectors expressing tagged dCas9 and guide RNA. The retroviral enChIP...plasmids for enChIP through Addgene. Retroviral Expression System for enChIP using CRISPR Addgene: Your ...enChIP system enables us to express the CRISPR complex much easier in various cells, including primary mouse... IFNgamma-Responsive Promoter by a Retroviral Expression System for enChIP Using CRISPR. Fujita T, Fujii...

  17. CRISPR Between the Genes: How to Experiment with Enhancers and Epigenomics

    Type
    Blog Post
    Published
    Aug. 9, 2016, 2:30 p.m.
    ... DNA compaction, an enhancer can increase the expression of a gene that is millions of base pairs away...Putting it all together, we get the following expression for step 1: (H3K4me1 + H3K27ac) – H3K4me3 The...enhancers with CRISPR Armed with the genome math expression for NANOG enhancers, we’re ready for step 2: ...of embryonic stem cells (ESCs) with a lentiviral vector, an attenuated retrovirus in the same family as...regions and their reverse complements were parsed for spCas9 PAM sites (NGG) and then filtered based on two ...
Showing: 101 - 117 of 117 results