Assess CUT&RUN-enriched DNA

Analyze CUT&RUN-enriched DNA prior to library prep. Consider the following:

DNA yield

There is no typical DNA yield for CUT&RUN, as yields can vary by cell type, number of cells, target abundance, and antibody quality. Instead, we suggest to:

  • Check that yields from the H3K4me3 control are similar to or slightly greater than the IgG negative control.
    • Note that similar and/or low yields from the positive control reflect the low abundance of H3K4me3 and do NOT imply assay failure. Read more here.
    • If a positive control with higher yields is needed, we recommend H3K27me3 (EpiCypher 13-0055).
  • Aim for ≥5ng CUT&RUN-enriched DNA, which will enable robust library prep.
    • Yields below 5 ng are common for low abundance targets, such as H3K4me3. In these cases, we suggest using all of the CUT&RUN DNA for library prep.
    • For help with low yields, see this FAQ. Note that sometimes low yields cannot be avoided, but modifications can be made to library prep to increase yields for sequencing.

Fragment analysis

Do NOT assess fragment size distribution of CUT&RUN DNA. Raw CUT&RUN yields are too low for detection on Bioanalyzer/TapeStation, and will not provide useful information at this step. Wait until after library prep. For further reading, see this FAQ.

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