Question: How do I measure gDNA fragment length?
Answer: We recommend the following methods for size measurement of high-molecular weight (HMW) genomic DNA:
- Pulsed field gel electrophoresis (i.e. CHEF PFGE system from Bio-Rad): Most accurate method for sizing DNA >50 kb. This method can determine DNA size up to 5 Mb. It requires a minimum of 100 - 200 ng input DNA.
- Pippin Pulse (from Sage Science): This method is not as accurate as PFGE but can roughly approximate the percentage of molecules >50 kb. It has reduced resolution for large (>50 kb) DNA molecules and can have run-to-run variability. However, it can be useful as a screening tool for degraded samples. It requires a minimum of 100 - 200 ng input DNA.
- Capillary-based methods (i.e. Agilent TapeStation, Caliper LabChip GX, Advanced Analytical Fragment Analyzer): These methods are not as accurate as either PFGE or Pippin Pulse for measuring large DNA fragments but can roughly approximate the percentage of molecules >50 kb. The sizing accuracy declines for molecules >50 kb. However, it can be useful as a screening tool for degraded samples. It requires a minimum of 10 - 20 ng input DNA.
Note: The concentration of DNA in samples analyzed by these methods should be > 10ng/ul. Standard agarose gels are not recommended for determining size distribution of HMW DNA.
For more information, see the Demonstrated Protocol on High Molecular Weight DNA QC: https://support.10xgenomics.com/genome-exome/index/doc/demonstrated-protocol-hmw-dna-qc
Products: Genome, Exome