Temperature-dependence of preconditioning for lengthened capillary DNA sequencing

Gary A. Griess, Stephen C. Hardies, Philip Serwer

Research output: Contribution to journalArticlepeer-review

Abstract

A previous study shows that electrophoretic preconditioning of a commercial polymer solution increases the spacing and resolution of DNA fragments fractionated in this solution by CE at 50°C (Griess, G. A. et al., Electrophoresis 2005, 26, 102). The present study shows that this preconditioning effect on peak spacing progressively increases when the temperature of preconditioning increases to 70°C, though fractionation is still performed at 50°C. An increase in peak sharpness accompanies the increase in peak separation for DNA fragments longer than ∼200 bases. Changing the preconditioning temperature from 50 to 70°C optimally improves resolution of fragment analysis in the range of 600-2000 nucleotides. When DNA sequencing is performed with automated base calling and 70°C preconditioning at 319 V/cm (47 cm long capillary, Applied Biosystems 310 apparatus), the range of high-quality base calls is increased by 25% to ∼750; the range of low-quality base calls is increased by about 100% to ∼1200 in comparison to DNA sequencing without preconditioning.

Original languageEnglish (US)
Pages (from-to)4440-4448
Number of pages9
JournalELECTROPHORESIS
Volume26
Issue number23
DOIs
StatePublished - Dec 2005

Keywords

  • Capillary electrophoresis
  • Genomics
  • Peak sharpening
  • Polymer solution
  • Sieving

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Clinical Biochemistry

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