DNA Fragmentation (E2600)

Protocol

1. DNA may be fragmented using sonication, nebulization, enzymatic treatment or by other methods. Fragments must average less than 1,000 bp and should be in DNase-free TE buffer (pH 7.5). Determine the approximate size distribution of the DNA by agarose gel electrophoresis of the sample alongside a DNA marker (e.g., 50 bp DNA Ladder, NEB #N3236 ). A sample of fragmented HeLa genomic DNA of the size range of 100–1000 bp is included as a control. 4 μl of this DNA standard (~400 ng) is sufficient to visualize on an agarose gel alongside the experimental sample. The desired fragment size should be appropriate for the desired downstream analysis. For example, DNA fragmented to an average length of ~250 bp is suitable for assay by real-time quantitative PCR (qPCR). Smaller fragments of 150 bp or less are suitable for linker addition in Next Generation Sequencing platforms. It is also important to quantitate the amount of DNA in the experimental sample by A₂₆₀ measurement using a spectrophotometer, Nanodrop™ instrument or Bioanalyser™.