A1: A comparison of the sequencing results between genomic DNA prepared with NEBNext dsDNA Fragmentase and with mechanical shearing demonstrates that the NEBNext dsDNA Fragmentase does not introduce any detectable bias during the sequencing library preparation and no difference in sequence coverage is observed between the two methods.
Q2: When do I need to use the E.coli ligase for Fragmentase?
A2: If the DNA fragments created by dsDNA Fragmenase will be exposed to a strand-displacing enzyme such as Klenow exo-, before being treated with a ligase, then E. coli DNA Ligase for Fragmentase should be used. If the fragmented DNA will be used for sample preparation for Illumina sequencing then the E. coli DNA Ligase for Fragmentase should be used.
Q3: Can I add E. coli DNA Ligase for Fragmentase to the Fragmentase reaction?
A3: No, the E. coli DNA Ligase for Fragmentase must be used after the Fragmentase reaction has been stopped.
Q4: Can I add E. coli DNA Ligase for Fragmentase to my end repair reaction?
A4: Yes. Addition of E. coli DNA Ligase for Fragmentase to standard end repair reactions under standard conditions will be effective.
Q5: What sequencing platforms can DNA fragments produced using dsDNA Fragmentase be used for?
A5: DNA produced by dsDNA Fragmentase can be used for Illumina, 454 and SOLiD sequencing.
Q6: Can dsDNA Fragmentase be used with GC-rich or AT-rich DNA?
A6: Yes. The incubation times for GC-rich and AT-rich DNA are slightly different and these are described in the dsDNA Fragmentase protocols on the NEB website. Sequencing of GC-rich and AT-rich DNA prepared using dsDNA Fragmentase or Covaris shearing was found to be equivalent.
Q7: Can dsDNA Fragmentase be used with PCR and RT-PCR products?
A7: Yes. This is described in the dsDNA Fragmentase protocol link.
Q8: Can dsDNA Fragmentase be used with Whole Genome Amplified DNA (WGA DNA)?
A8: Yes. This is described in the dsDNA Fragmentase protocol link.
