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Showing: 1 - 20 of 34 results

  1. Genomic Deletions in Mammalian Cell Lines

    Type
    Collection
    ...to perform PCR in step 6.3 . Assemble a 20 μl PCR with the following components: 10 μl 2x PCR mix, 0.5 ...each well) to a 96-well PCR plate using a multichannel pipette. Centrifuge PCR plate at 400 x g for 5 ... in prostate tumors by quantitative real-time PCR (qPCR). Gordanpour A, Nam RK, Sugar L, Bacopulos...biallelic modification, ease of rapid identification by PCR, predictability of loss-of-function, and utility ... genomic DNA (gDNA) for the intended deletion by PCR (see steps 6.1 and 7.1 ). The bulk cells have sufficiently...sgRNA-B (see step 5 ). The purpose of the following PCR is to validate primers and verify the presence of...Use the primers designed in step 2 above. Conduct PCR for “non-deletion band” and “deletion band” in separate...

  2. Plan Your Experiment

    Type
    Collection
    ...cell population. The region of interest is PCR amplified, PCR products are denatured-renatured, treated... The PCR product is then run on an agarose gel to determine whether the edit was successful. PCR amplification...run on an agarose gel to identify DNA fragments. PCR and restriction digest (for HDR repaired DSBs): For...novel restriction site. The region of interest is PCR amplified, digested with the appropriate restriction...run on an agarose gel to identify DNA fragments. PCR amplification and gel electrophoresis (for HDR or...deletions or insertions, the region of interest can be PCR amplified using primers that (A) flank the region...and exact sequence of targeted alleles. Involves PCR amplification of targeted region from DNA, subcloning...

  3. Viral Production

    Type
    Collection
    ... AAV particles are titered by droplet digital PCR (ddPCR) using primers and probes targeting the ITR elements...April 2016) also performed by droplet digital PCR (ddPCR). Quality Control Addgene ensures high quality...are titered using a lentiviral droplet digital PCR (ddPCR) titration protocol (modified from Wang et al...confirmed to have the correct integrated provirus using PCR. This assay involves transducing cells with serial...obtained using ddPCR are generally 2–3 fold higher than those achieved using the standard qPCR method. Titering...

  5. Zhang Lab's CRISPR Frequently Asked Questions

    Type
    Collection
    ...better for PCR and surveyor. I used DNA polymerase Takara Ex Taq ™ DNA Polymerase for my genomic PCR, but couldn't...conditions with 5% DMSO or without 5% DMSO. My genomic PCR didn’t work, do you have any suggestions? The Zhang...the protocol, then uses around 2-4ul in each 50ul PCR reaction. The gDNA extracted in this way is usually...groups have successfully used Herc II or Kapa Hifi in PCR. Maybe this Takara enzyme is not very robust in this...

  6. CRISPR Plasmids - Tagging

    Type
    Collection
    ... technique for use in Drosophila cells that uses PCR to generate both an expression cassette for the Cas9.... The plasmids in the following articles provide PCR templates for amplification of the tag (eg GFP, Flag...expresses Cas9, PITCh-gRNA, & FBL-specific gRNA pCRIS-PITChv2-FBL - PITCh donor vector for EGFP-2A-PuroR...

  7. Zhang Lab CRISPR Page

    Type
    Collection
    ... be co-transfected as PCR amplicons using a protocol similar to SpCas9 sgRNA PCR (Ran et al., Nature Protocols...codon-optimized SpCas9; sgRNA can be co-transfected as PCR amplicons (Ran et al., Nature Protocols 2013). Individual...

  8. CRISPR Plasmids - C. elegans

    Type
    Collection
    ...need Cas9 plasmid Joung 46170 PU6::klp-12_sgRNA cU6 PCR (see paper) Transfection none, need Cas9 plasmid ...

  9. Jaenisch Lab CRISPR Plasmids

    Type
    Collection
    ...containing the sgRNA expressing fragment produced by PCR), or expressed from a single dual-expression vector...48214 pAC1-pCR8-dCas9VP48 : dCas9VP48 on Gateway donor vector pCR8/GW/TOPO. 48218 pAC84-pCR8-dCas9 : dCas9...donor vector pCR8/GW/TOPO. 48219 pAC147-pCR8-dCas9VP64 : dCas9VP64 on Gateway donor vector pCR8/GW/TOPO. ...48220 pAC148-pCR8-dCas9VP96 : dCas9VP96 on Gateway donor vector pCR8/GW/TOPO. 48221 pAC149-pCR8-dCas9VP160...dCas9VP160 : dCas9VP160 on Gateway donor vector pCR8/GW/TOPO. Table 4. Gateway Destination Vector ID Plasmid ...

  11. CRISPR Plasmids - Empty gRNA Vectors

    Type
    Collection
    ... none S. pyogenes Kamoun pUC119-gRNA 52255 Plant PCR template none S. pyogenes Sheen pRGEB31 51295 Plant...none S. pyogenes Boxem PU6::klp-12_sgRNA 46170 Worm PCR (see paper) none S. pyogenes Calarco SP6-sgRNA-scaffold...pyogenes Sternberg pJW1311 61254 Worm none, fusion PCR none S. pyogenes Ward pCFD1-dU6:1gRNA 49408 Fly BbsI...Genome Editing in Monocot and Dicot Plants Plant PCR-based procedure to rapidly generate multiple sgRNA... pyogenes Conklin pcrRNA.ind 61284 Bacteria araB none S. pyogenes Beisel pcrRNA.con 61285 Bacteria J23119...Enzyme Co-expressed Cas9 Cas9 System Selection PI pCRISPR 42875 Bacteria BsaI none S. pyogenes Chloramphenicol...42250 Zebrafish BsaI none S. pyogenes Joung pCRISPathBrick 65006 Bacteria BsaI yes, interfere S. pyogenes...

  13. Antibody Production

    Type
    Collection
    ...heavy chain and/or light chain variable regions. The PCR product is purified, sequenced via the Sanger method...

  14. CRISPR Plasmids - Plants

    Type
    Collection
    ...BsaI none S. pyogenes Kamoun 52255 pUC119-gRNA U6 PCR template none S. pyogenes Sheen 51295 pRGEB31 rice...

  15. Open Enzyme Collection

    Type
    Collection
    ...this collection include polymerase chain reaction (PCR), cloning, isothermal amplification, reverse transcription...

  16. TALEN Plasmids and Kits

    Type
    Collection
    ...somatic cells. Keith Joung Zhang Lab TALE Toolbox PCR/Golden Gate cloning method. Optimized for human expression...

  17. CRISPR References and Information

    Type
    Collection
    ...the tool, you'll need Sanger sequencing reads from PCR amplicons that cover your locus of interest and correspond...PDF 125.6 KB Marraffini Bacteria: pCRISPR new spacer cloning pCRISPR PDF 106.7 KB Mendenhall & Myers Mammalian...

  18. Retrograde AAV viral preps

    Type
    Collection
    ...AAV titering by SYBR green qPCR , confirmation of the transfer plasmid with PCR, purity assessment by silver...

  19. Caltech Systemic Capsids

    Type
    Collection
    ...including AAV titering by probe-based droplet digital PCR, in vitro and (when possible) in vivo testing, and...

  20. Cre-lox system

    Type
    Collection
    ...pBS500 EF1alpha-GFPcre Cre-GFP fusion EF-1 alpha Mammalian Sauer 11923 pBS598 EF1alpha-EGFPcre Cre-EGFP fusion...EF1alpha-EGFPcre* Cre-EGFP fusion EF-1 alpha Mammalian Sauer 11956 pBS594 promoterless EGFPcre Cre-EGFP...tet-hCMV-EGFPcre Cre-EGFP fusion Tet inducible Mammalian Sauer 11958 pBS532 promoterless GFPcre promoterless...-hCMV-GFPcre tet inducible Cre-GFP fusion tet-hCMV Mammalian Sauer 11961 pBS596 tet-hCMV-GFPcre tet inducible...Cre EF-1 alpha Mammalian Sauer 11919 pBS448 RSV-GFPcre Cre-GFP fusion Rous sarcoma virus (RSV) LTR/promoter...Tamoxifen-inducible Cre None Zebrafish Stankunas 82696 pCRE-iRFP670 Cre and iRFP670 PGK Mammalian Mullen 84032...
Showing: 1 - 20 of 34 results