Advantages of using the QIAshredder instead of restriction digestion to prepare DNA for droplet digital PCR

Steven A. Yukl, Philip Keiser, Peggy Kim, Peilin Li, Joseph K. Wong

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

The viscosity of genomic DNA can interfere with digital PCR systems that partition samples into oil droplets or microfluidic wells. Restriction digestion may reduce the viscosity, but the process is labor-intensive, and the buffer can alter the conditions for PCR. DNA fragmentation using the QIAshredder (a biopolymer spin column) is faster, may result in more predictable and uniformly-sized fragments, and avoids the need for restriction buffers that can inhibit downstream PCR. In 10 separate head-to-head experiments comparing aliquots of DNA processed using the QIAshredder to those digested with RsaI or BsaJI prior to droplet digital PCR, we found that the copy numbers measured from the QIAshredded DNA tended to be greater than those measured from the digested DNA (average of 1.35-fold compared with BsaJI; P <0.0001), even for inputs as high as 1.8 μg or dilution down to the single copy level.

Original languageEnglish (US)
Pages (from-to)194-196
Number of pages3
JournalBioTechniques
Volume56
Issue number4
DOIs
StatePublished - 2014
Externally publishedYes

Fingerprint

Digestion
Polymerase Chain Reaction
DNA
Viscosity
Buffers
Biopolymers
Microfluidics
DNA Fragmentation
Oils
Dilution
Personnel
Experiments

Keywords

  • BsaJI
  • Digital PCR
  • DNA
  • Droplet Digital PCR
  • QIAshredder
  • Restriction enzyme
  • RsaI

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Biotechnology
  • Medicine(all)

Cite this

Advantages of using the QIAshredder instead of restriction digestion to prepare DNA for droplet digital PCR. / Yukl, Steven A.; Keiser, Philip; Kim, Peggy; Li, Peilin; Wong, Joseph K.

In: BioTechniques, Vol. 56, No. 4, 2014, p. 194-196.

Research output: Contribution to journalArticle

Yukl, Steven A. ; Keiser, Philip ; Kim, Peggy ; Li, Peilin ; Wong, Joseph K. / Advantages of using the QIAshredder instead of restriction digestion to prepare DNA for droplet digital PCR. In: BioTechniques. 2014 ; Vol. 56, No. 4. pp. 194-196.
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