asCpf1 Nuclease NLS Protein (250 pmol)

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K08838 μg (250 pmol/ 25µL)
$70.00

Customize asCpf1 Nuclease NLS Protein (250 pmol)
1 x asCpf1 Nuclease NLS Protein   + $0.00
1 x 10X Reaction Buffer   + $0.00

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asCpf1 Nuclease NLS Protein (250 pmol)
asCpf1 Nuclease NLS Protein (250 pmol)

In stock

$70.00

Summary
    Specifications


    Description

    Using Cpf1 (a.k.a. Cas12a) in your CRISPR experiment offers several advantages over other CRISPR-associated nucleases.

      • Due to the T-rich PAM sequence (TTTN), Cpf1 enables editing in regions unable to be targeted by Cas9.
      • Cpf1 can be used with a shorter guide RNA (called crRNA) than Cas9.
      • Cpf1 creates a staggered cut in dsDNA instead of a blunt cut.
      • Cpf1 cuts distal to the PAM sequence, which may allow for multiple rounds of cleavage.
        • asCpf1 is from the bacteria

    Acidaminococcus

        . This protein contains a SV40 T antigen nuclear localization signal (NLS) on the N-terminus of the protein. If the cut caused by asCpf1 is repaired by non-homologous end joining (NHEJ), an indel may be formed that disrupts the open reading frame of the targeted gene, leading to gene knockout. Alternatively, by supplying a repair template, a sequence can be knocked in at the cleavage site via homology directed repair (HDR).
    SKUK088
    Cas TypeNuclease
    Cas OriginasCpf1
    Cas Protein MarkerNo GFP
    Source

    E. coli

    AliasCas12a, asCas12a, Cpf1 from Acidaminococcus, Cas12a from Acidaminococcus
    Unit quantity38 μg (250 pmol/ 25µL)
    Product Concentration10 μM, 1.5 mg/ml
    FormatEnzyme supplied with 10X Reaction Buffer.
    Storage Buffer10 mM Tris-HCl (pH 7.4), 0.1 mM EDTA, 1 mM DTT, 300 mM NaCl, and 50% (v/v) Glycerol.
    StorageStore all components at -20°C.
    CautionThis product is distributed for laboratory research only. Caution: Not for diagnostic use.
    Documents


    Supporting Protocol
    MSDS

      QC

        Other
        FAQs


        What is the in vitro digestion of DNA protocol with Cas9 Nucleases?
        1. 1. Mix individual components before use and assemble reaction at room temperature.
          Component Volume
          sgRNA (300 nM) 3 µl
          Cas9 Nuclease Protein (1 µM) 1 µl
          10X Cas9 Reaction Buffer 3 µl
          Nuclease-free H2O 20 µl
          Pre-incubate without substrate for 15 min at 37°C
          Substrate DNA (30 nM) 3 µl
          Total Volume 30 µl

        2. 2. Mix thoroughly, and pulse-spin in a microfuge.
        3. 3. Incubate at 37°C for 1 hour.
        4. 4. (Optional) Add 1 μl of Proteinase K to each sample. Mix thoroughly and pulse-spin in a microfuge. Incubate at room temperature for 10 minutes.
        5. 5. Analyze fragments via agarose gel electrophoresis.

        The substrate DNA: sgRNA : Cas9 molar ratio must be kept at 1:10:10 for highest efficiency.
        References


        1
        • Nelson, C. E., Wu, Y., Gemberling, M. P., Oliver, M. L., Waller, M. A., Bohning, J. D., … Gersbach, C. A. "Long-term evaluation of AAV-CRISPR genome editing for Duchenne muscular dystrophy" Nature Medicine 25(3):427–432 (2019). DOI: 10.1038/s41591-019-0344-3.