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High Throughput PAM Determination Assay Libraries
(Pooled Libraries #160132, #160133, #160134, #160135)

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  • Purpose

    These pooled libraries encode CRISPR-Cas target sites flanked by randomized nucleotides in place of a protospacer adjacent motif (PAM). The libraries are used to determine the PAM requirements of Cas enzymes.

    Each library differs in the CRISPR-Cas target site sequence, the orientation of the randomized nucleotide sequence, and the library complexity.

    Libraries
    • Library 160132 — p11-3’_8xN_PAM-site1 (RTW554)
      • For characterizing Cas9 PAMs
      • Randomized 8 nt PAM 3' of target site 1
    • Library 160133 — p11-3’_8xN_PAM-site2 (RTW555)
      • For characterizing Cas9 PAMs
      • Randomized 8 nt PAM 3' of target site 2
    • Library 160134 — p11-5’_10xN_PAM-site3 (RTW572)
      • For characterizing Cas12a PAMs
      • Randomized 10 nt PAM 5' of target site 3
    • Library 160135 — p11-5’_10xN_PAM-site4 (RTW574)
      • For characterizing Cas12a PAMs
      • Randomized 10 nt PAM 5' of target site 4
  • Vector Backbone

    p11-LacY-wtx1 (Plasmid #69056)

Ordering

Item Catalog # Description Quantity Price (USD)
Pooled Library 160132 p11-3'_8xN_PAM-site1 1 $325 Add to Cart
Pooled Library 160133 p11-3'_8xN_PAM-site2 1 $325 Add to Cart
Pooled Library 160134 p11-5'_10xN_PAM-site3 1 Pending
Pooled Library 160135 p11-5'_10xN_PAM-site4 1 $325 Add to Cart
Available to Academic and Nonprofits Only

Library Details

Please see the Purpose section for library details.

Library Shipping

This library is delivered as suspended DNA in a microcentrifuge tube on blue ice. The tube's contents will not necessarily be frozen. For best results, minimize freeze/thaws.

  • Volume
    ∼10 µL
  • Concentration
    1 ng/µL

Resource Information

Depositor Comments

The Kleinstiver Lab created a Github repository sharing the code to perform an analysis of data generated using the HT-PAMDA method.

How to cite this pooled library ( Back to top )

These pooled libraries were created by your colleagues. Please acknowledge the Principal Investigator, cite the article in which the plasmids were described, and include Addgene in the Materials and Methods of your future publications.

  • For your Materials & Methods section:

    High Throughput PAM Determination Assay p11-3'_8xN_PAM-site1 Library was a gift from Keith Joung and Benjamin Kleinstiver (Addgene #160132)
    High Throughput PAM Determination Assay p11-3'_8xN_PAM-site2 Library was a gift from Keith Joung and Benjamin Kleinstiver (Addgene #160133)
    High Throughput PAM Determination Assay p11-5'_10xN_PAM-site3 Library was a gift from Keith Joung and Benjamin Kleinstiver (Addgene #160134)
    High Throughput PAM Determination Assay p11-5'_10xN_PAM-site4 Library was a gift from Keith Joung and Benjamin Kleinstiver (Addgene #160135)

  • For your References section:

    Unconstrained genome targeting with near-PAMless engineered CRISPR-Cas9 variants. Walton RT, Christie KA, Whittaker MN, Kleinstiver BP. Science. 2020 Apr 17;368(6488):290-296. doi: 10.1126/science.aba8853. Epub 2020 Mar 26. PubMed 32217751
    Scalable characterization of the PAM requirements of CRISPR-Cas enzymes using HT-PAMDA. Walton RT, Hsu JY, Joung JK, Kleinstiver BP. Nat Protoc. 2021 Feb 5. pii: 10.1038/s41596-020-00465-2. doi: 10.1038/s41596-020-00465-2. PubMed 33547443
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