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We narrowed to 716 results for: cas

Showing: 201 - 220 of 716 results

  1. Pouring LB Agar Plates

    Type
    Protocol
    ...consisting of: 5.0 g yeast extract 10.0 g peptone from casein 10.0 g sodium chloride 12.0 g agar-agar 1 L Sterile...actually make a bit more than 200 mL (~220 mL) just in case we spill anything or have small errors in measurement...the autoclave. Transfer the sterile water (in our case 220 mL) to the same bottle and swirl to form a medium...s manufacturer to make sure this is true in your case. Retrieve your molten agar mix from the autoclave...positive and negative test results. Sample Data In all cases below (-) indicates that the tested strain is not...

  2. Western Blot

    Type
    Protocol
    ...protein concentration in the lysate, running a precast SDS-PAGE gel, and immunoblotting....protein concentration in the lysate, running a precast SDS-PAGE gel, and immunoblotting. Sharing speeds...23227 β-mercaptoethanol 4X protein loading buffer Precast SDS-PAGE gel SDS-PAGE running buffer Prestained...min at 100 °C . Section 3: SDS-PAGE Prepare the precast gel as follows: Remove the gel from the plastic...

  3. Plasmids 101: Using Transposons in the Lab

    Type
    Blog Post
    ...in Vibrio cholerae that encodes a Type I-F CRISPR-Cas system. This system has four major components: 1)...crRNA), 2) four proteins (TniQ, Cas8, Cas7, Cas6) that forms the QCascade DNA-targeting module with the ... transposons encode reporter genes, mutagenic cassettes, or barcodes. When transposons are delivered to...For the INTEGRATE system to transpose, first the QCascade complex surveys the genome and binds its target...molecule, and inserts it ~50 bp downstream of QCascade’s genomic target site (Fig. 3B). The INTEGRATE ...of RNA-guided transposition with INTEGRATE. The QCascade complex surveys the genome and binds its target...molecule, and inserts it ~50 bp downstream of QCascade’s genomic target site.  T-RL= transposon right-...

  4. Viral Vectors 101: Optogenetic Tools

    Type
    Blog Post
    ...allow positively charged ions such as Na+, K+, and Ca++ to passively enter the cell with the concentration...function through multiple intra-cellular signaling cascades. Optical methods for controlling GPCR activation...control GPCR activation and downstream signaling cascades (reviewed in Abreu & Levitz, 2020), including ...extracellular domain which activates the native signaling cascade of the intracellular part of the receptor, similar... out important branch points along a signaling cascade that may go awry in a disease state.      ...

  5. Ligation Independent Cloning

    Type
    Protocol
    ...and shift its activity back to polymerase. In our case, dGTP will be used in the reaction, meaning that...there are no gaps in the final assembly. In this case, 15 bp of homologous sequence is used, plus a minimum...following the manufacturer's instructions. In this case, we will include the free nucleotide dGTP in the...

  6. Colony Formation Titering Assay

    Type
    Protocol
    ...seeded the day before transduction. Note that in that case, polybrene should only be added at the time of viral...sensitive to lentivirus or to polybrene, in which case this incubation step can be reduced to 8 h and the...serial dilutions of the lentiviral vector pHAGE EF1α dCas9-KRAB . The following day, cells were treated with...

  7. Optogenetics Guide

    Type
    Guide
    ...Methods. 9(2):159-72. PMID 22179551 Saunders A, Johnson CA, Sabatini BL 2012. Novel recombinant adeno-associated...thus expressed in all GABAergic neurons. In this case, the subpopulation of GABAergic neurons being activated...

  8. Virus Protocol - Generating Stable Cell Lines

    Type
    Protocol
    ...A549 cells stably expressing Cas9. A549 cells were transduced with lentiCas9-Blast and then selected with...formed by expansion of single cells for 13 days. lentiCas9-Blast was a gift from Feng Zhang (Addgene plasmid...

  10. CRISPR 101: Multiplex Expression of gRNAs

    Type
    Blog Post
    ...Addgene. 10. Port, Fillip, et al. “Optimized CRISPR/Cas tools for efficient germline and somatic genome engineering...humanized Cas9, D10A nickase mutant (Cas9n), dCas9 (transcriptional repression), and Fok1-dCas9 (dimeric...options include humanized wt Cas9, dCas9 (transcriptional repressor), and dCas9-VP64 (transcriptional activator...including Standard PolIII-gRNA cassette, Csy4-cleavable cassette, and PTG cassette.    Find gRNA multiplexing... then digested using BsmBI and ligated into Cas9 or dCas9-containing destination vectors. Destination ... encoding the gRNAs and Cas9. As with the Gersbach lab plasmids, multiple Cas9 variants are available:...the multiple gRNAs and Cas9 together into a single plasmid. Although only wt hCas9 entry vectors are supplied...

  11. Protocol - How to Run an Agarose Gel

    Type
    Protocol
    ... how to perform gel electrophoresis. Equipment Casting tray Well combs Voltage source Gel box UV light...prefer to microwave in pulses, swirling the flask occasionally as the solution heats up.). Caution HOT! Be ...

  12. Plasmid Cloning by PCR (with Protocols)

    Type
    Protocol
    ...of a hairpin structure within your primer. In our case, we will add TAAGCA, resulting in a final Forward...to the sequence to be amplified (the ORF in this case), not the Tm of the entire primer. If you are amplifying...

  13. Antibody Validation Using the Indirect ELISA Method

    Type
    Protocol
    ...antibody concentration may have been too low. In this case, use higher antibody concentrations. If the reaction...antibody concentration may have been too high. In this case, use lower antibody concentrations. The ideal range...

  14. Lentiviral Prep Service

    Type
    Collection
    ...Root Cas9 Lentivirus Cas9 viruses can be used to interrogate the genome in a variety of ways. Cas9 proteins... 52962 lentiCas9-Blast Sp Cas9 Cut Blasticidin Expresses human codon-optimized S. pyogenes Cas9 protein...backbone. Zhang Cas9 and Accessories for Activating Gene Expression Catalytically-dead Cas9 (dCas9) can be fused...Resistance Activity PI 61422 dCAS9-VP64_GFP dCAS9 (D10A, H840A) none Expresses dCAS9-VP64 activator with 2A ...lenti dCAS-VP64_Blast dCAS9 (D10A, N863A) Blasticidin 3rd generation lenti vector encoding dCAS9-VP64 ...-targeting controls. This backbone contains SpCas9. SpCas9 and 76,441 unique sgRNAs targeting 19,114 human...plasmids resource page . Cas9 for Editing Genomic Sequences To perform genome edits, Cas9 nuclease is used to...

  15. DNA Quantification

    Type
    Protocol
    ...light that your substance maximally absorbs. In the case of nucleic acid (DNA and RNA), the maximal absorbance...

  16. Trimmer Lab NeuroMab Collection

    Type
    Collection
    ...Anti-CASPR/Neurexin IV [K65A/2R] CASPR/Neurexin IV Mouse IgG2a 206606 Anti-CASPR2 [K67/11R] CASPR2 Human...190491 CASK scFv [K56A/50] K56A/50 scFv CASK Human Mouse 190492 CASK scFv [K56A/57] K56A/57 scFv CASK Human...177445 Anti-CASPR/Neurexin IV [K65/35R] CASPR/Neurexin IV Rat Mouse IgG2a 177446 Anti-CASPR2 [K67/25R] ...219448 Anti-CASPR/Neurexin IV [K65A/30R] CASPR/Neurexin IV Rat Mouse IgG2a 219449 Anti-CASPR/Neurexin IV...SAP97 Rat Mouse IgG2a 128625 Anti-CASPR/Neurexin IV [K66/38R] CASPR/Neurexin IV Rat Mouse IgG2a 128626...Pan-KChIP K+ channel Mouse IgG2a 177443 Anti-CASK [K56A/50R] CASK Human Mouse IgG2a 177444 Anti-KChIP2b K+...Synaptotagmin-6 Mouse Mouse IgG2a 188217 Anti-CASK [K56A/57R] CASK Human Mouse IgG2a 188218 Anti-Kv2.1 K+ channel...

  17. Brzezinski Lab CRISPR Collection

    Type
    Collection
    ... for shuttling U6-guide cassettes to make dual guide expressing plasmids dCas9-KRAB-MeCP2 fusion for gene...enhancer sequence repression dCas9-VPR fusion for gene activation CRISPR/Cas9 Plasmid Variants Figure 1:...175574 express dCas9-KRAB-MeCP2 for repression. Plasmids 175571 and 175572 express dCas9-VPR for activation...in retinal development the lab utilizes a CRISPR/Cas9 based approach to knockout, repress, and activate...taken advantage of the Zhang lab all-in-one (guide/Cas9/reporter) pX458 plasmid family. They have made several...plasmids to insert a second guide within a U6-guide cassette. This process is compatible with all pX458 family...Sources Original pX458 backbone ( Addgene #48138 ) dCas9 + KRAB-MeCP2 ( Addgene #110821 ) VPR ( Addgene #63798...

  18. Video Library

    Type
    Protocol
    ... deletions in mammalian cells lines using CRISPR/Cas9 Genomic Deletions with CRISPR Protocol Over-Agar...

  19. Centrifugation

    Type
    Protocol
    ... not dislodge any pellets or re-mix liquids. In cases where you are separating solids from a liquid, pellets...

  20. Tetracycline Inducible Expression

    Type
    Collection
    ...Description PI 63800 PB-TRE-dCas9-VPR Tet-inducible expression of SP-dCas9-VPR transcriptional activator...tet-inducible Cas9-2A-EGFP expression. Based on LentiCRISPR v2 . Adam Karpf 167935 pLenti-tetON-KRAB-dCas9-DHFR-EF1a-TagRFP...Julien Béthune 154939 dCas13d-dsRBD-APEX2 RNA-protein interaction mapping via Cas13-based APEX targeting...your gene of interest should only be expressed occasionally, use Tet-On: express an rtTA or TetR and include...should be generally active and only turned off occasionally, use Tet-Off: express a tTA and include a tet-responsive...DHFR-EF1a-TagRFP-2A-tet3G Tet-inducible expression of KRAB-dCas9-DHFR for CRISPRi. See Plasmid #167937 for CRISPRa...Dox-inducible expression of Cre recombinase. Madeline Lancaster 198752 DiLiCre 2.0 Dox-inducible expression of...
Showing: 201 - 220 of 716 results