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We narrowed to 347 results for: ATC

Showing: 41 - 60 of 347 results

  1. Screening for Successful Genome Editing with Digital PCR

    Type
    Blog Post
    ...projects. Most researchers instead turn to so-called “mismatch nuclease” assays (e.g. Surveyor® or T7E1) for ...technology. Digital PCR has several advantages over mismatch nuclease assays that will be elaborated below....several major advantages over the commonly employed mismatch nuclease assays. Sensitivity: Less than 20 ng...plate without prior quantification. Comparatively, mismatch assays require between 200 and 500 ng of purified...about 5% mutant targets have been reported for mismatch nuclease assays (2, 6, 7). Ability to distinguish...complete functional knockout of your target gene. Mismatch nuclease assays are completely “blind” to this.... & Pollet, N. Comparison of T7E1 and surveyor mismatch cleavage assays to detect mutations triggered ...

  2. Cpf1 Update: Comparison to Cas9 and NgAgo

    Type
    Blog Post
    ...pair gRNA-target mismatches on modification by AsCpf1 (red) or LbCpf1 (blue). Mismatch positions are indicated...with single and double mismatches throughout the 23-base sequence. Double mismatches ablated Cpf1 activity...bases 19-23). Cpf1 is also sensitive to single mismatches, but variably so, with Kleinstiver et al. reporting...reporting that Cpf1 can tolerate mismatches at gRNA positions 1, 8, 9, and 19-23. Accordingly, the 3’ end...PubMed PMID: 26411297 Resources on the Addgene Blog Catch Up On Your Cpf1 Background Read About CRISPR Competitor...

  3. Controlling for Off-target Effects with a New Genome-wide CRISPR Screen Design

    Type
    Blog Post
    ...exact or 1-bp mismatch off-targets had greater toxicity than guides that had zero mismatch off-targets....off-target activity: 1) mismatches closer to the PAM are less tolerated than mismatches more distal to the ...off-targets. Guides with 2-bp mismatch off-targets were only toxic if they had 5+ off-target sites. Results from...comparing the toxicity of guides that have 1-bp mismatch off-target sites, short guides had reduced toxicity...currently available from Addgene Other Resources Watch Dr. Michael Bassik’s lecture on “Multiplexing with...

  4. CRISPR Pooled gRNA Libraries

    Type
    Collection
    ...-Burgk 3rd 4 76,441 No available items found to match all requested criteria. Please consider using fewer...

  6. Antibodies 101: Conventional vs Spectral Flow Cytometry

    Type
    Blog Post
    ...antibodies are from the same batch, to avoid unmixing issues from batch-to-batch variation. Tandem dyes (two...conjugated together) can be especially sensitive to batch effect. For example, when a PE molecule is conjugated... Cy7 molecules are added during that particular batch. This can present particular difficulty in unmixing...sample is stained with a PE/Cy7 antibody from a batch that is not the same as your single color control...

  7. Cpf1: A New Tool for CRISPR Genome Editing

    Type
    Blog Post
    ...with single and double mismatches throughout the 23-base sequence. Double mismatches ablated Cpf1 activity...pair gRNA-target mismatches on modification by AsCpf1 (red) or LbCpf1 (blue). Mismatch positions are indicated...bases 19-23). Cpf1 is also sensitive to single mismatches, but variably so, with Kleinstiver et al. reporting...reporting that Cpf1 can tolerate mismatches at gRNA positions 1, 8, 9, and 19-23. Accordingly, the 3’ end...

  8. QC Sequencing Technologies at Addgene

    Type
    Blog Post
    ...there is a 100% nucleotide match to base identity and plasmid length. Any mismatch or indel flags the sequence...pipeline are now run through an auto-align process to match sequence results to full-length predicted sequences...differences. This script compares the sequence for a 100% match to base identity and length, but disregards the ...discrepancies. Any sequences that are a perfect match from this script can then continue in the auto-align...bioinformatic analysis. At Addgene, we use a mix-and-match approach for sequencing methods and choose the best...

  9. CRISPR 101: Off-Target Effects

    Type
    Blog Post
    ...“Off-target effects” a vague catch-all term for unintended consequences in an experiment. For CRISPR ...to select a Cas enzyme with low tolerance for mismatches within the gRNA sequence. Graphic courtesy... or mutated to be more specific for exact gRNA matches (Rabinowitz and Offen). Of note, most of these ...these optimized Cas enzymes prevent cutting at mismatched guide sites, but not necessarily binding to these... activity at those sites, especially when DNA mismatches are involved. Two other approaches have been ...non-targeting’ gRNA, a gRNA which doesn’t perfectly match any site in the genome. While this may effectively...

  10. Identifying Sequence Elements with SnapGene's Feature Database

    Type
    Blog Post
    ...devising rules for identifying a feature even when the match is imperfect. SnapGene's feature database The source...detection algorithm that tolerates occasional mismatches or indels. Empirical tests indicated that a reasonable...to allow searches for a perfect protein sequence match even when the DNA sequence has changed. Identifying...database. Another limitation is that by tolerating mismatches, our algorithm could annotate a feature inaccurately... strain to newly sequenced strains through our mismatch tolerant detection algorithm and thereby speed...

  11. Biosensor AAV Preps

    Type
    Collection
    ...Constitutive 1, 2 Li No available items found to match all requested criteria. Please consider using fewer...

  12. Neurodegeneration Plasmid Collection

    Type
    Collection
    ...Aebischer 10881 pCCL cPPT PGK EGFP WPRE LTR H1 shSOD1mismatch SOD1 H1 ALS Patrick Aebischer 11442 pBabe bleo...Mix HNRNPA1 His T7 ALS Tanja Mittag 169719 A1-LCD Patchy HNRNPA1 His T7 ALS Tanja Mittag 170443 pCDH-EF1a-hDNAJC6...neuropathy Albert La Spada No available items found to match all requested criteria. Please consider using fewer...

  13. SARS-CoV-2/COVID-19 Detection Methods Based on CRISPR/Cas

    Type
    Blog Post
    ...to the presence of mismatches within the DNA and the positions of these mismatches. To detect SARS-CoV...catalytically active in the presence of two or more mismatches in the cRNA:target duplex and will not give a...dCas9 complexes containing sgRNA that have no mismatches bind to the immobilised target sequences. Upon...making it an affinity-based method. Due to the mismatch sensitivity of FnCas9, this is also a good way...

  14. Tips for a 1st Time CRISPR User (by a 1st Time CRISPR User)

    Type
    Blog Post
    ...cleaves double stranded DNA (dsDNA) at basepair mismatches. To use this tool to detect a BRAF gene edit,... control cells, which should contain basepair mismatches at the site of the edit. Using Surveyor nuclease...nuclease followed by gel electrophoresis, these mismatches are indicated by the presence of DNA cleavage ...isn’t in the middle of the PCR product. If the mismatch is in the middle of the PCR product, you would...cells. Surveyor nuclease was used to digest (+) mismatches in DNA duplexes (513 bp), as indicated by the...

  15. Lambda Red: A Homologous Recombination-based Technique for Genetic Engineering

    Type
    Blog Post
    ...methyl -directed mismatch repair (MMR) system. MMR’s job is to correct DNA mismatches that occur during...activation.  The first way is to introduce a C/C mismatch at or within 6 base pairs of the edit site. This...when activation of MMR is avoided Introduce C/C mismatches or silent changes in wobble codons to avoid MMR...Enhanced levels of λ Red-mediated recombinants in mismatch repair mutants. Proceedings of the National Academy...

  16. SpyLigase Irreversibly Locks Peptides Together for Efficient Cell Capture

    Type
    Blog Post
    ..., just 13 amino acids, but SpyCatcher is a protein. You can fuse SpyCatcher onto things, but even better...particular. They recently introduced the SpyTag/SpyCatcher system, based on a protein isolated from Streptococcus...It came from our work in developing the SpyTag/SpyCatcher system, to be able to grab on to proteins irreversibly...

  17. Plasmids 101: How to Verify Your Plasmid Using a Restriction Digest Analysis

    Type
    Blog Post
    ...construct, and verify that the plasmid you received matches the expected sequence. Here at Addgene, we use ... 3) produces bands at 6kb and 1.2kb (red box), matching the backbone and insert, respectively. The results...results on the gel correspond to the predicted sizes. Watch this video for a quick overview of how to analyze...the optimal working conditions for each enzyme. Watch out for methylation issues. Enzymes like XbaI and...

  18. Plasmids 101: Methylation and Restriction Enzymes

    Type
    Blog Post
    ...role in regulating genome replication, repairing mismatched basepairs or small indels that occur during DNA... methylase adds a methyl group to the adenine of GATC stretches of DNA Dcm methylase adds a methyl group...and will therefore have methylated adenines in any GATC sequences found in the plasmid. When the PCR products...mutation (as these would also be deficient in mis-match repair functions of Dam), so these strains should...

  19. Career Coaching for Scientists: Why and Where Do I Find One?

    Type
    Blog Post
    ...out there, so finding a good one that is a style match for your needs can take some research.  It helps...coaches and I can't tell you which one will be a match for your needs, but here are a number I have come...group is that sometimes this offers the option of matching with the best person in the group for your needs... PhD is in History, I think she would be a good match as a coach for many scientists.  Sometimes it can...

  20. Design Tips for Prime Editing

    Type
    Blog Post
    ...bases. DNA mismatch repair (MMR) is less efficient at identifying these “bubbles” of mismatched bases, enhancing...efficiently targets single-base mismatches (left), but “bubbles” of 3 or more mismatched bases can often evade ...
Showing: 41 - 60 of 347 results