Parameters involved in the conversion of real-time PCR assays from the ABI prism 7700 to the Cepheid SmartCycler II

Heather A Bentley; Dorothy R Belloni; Gregory J Tsongalis

doi:10.1016/j.clinbiochem.2004.10.009

Parameters involved in the conversion of real-time PCR assays from the ABI prism 7700 to the Cepheid SmartCycler II

Clin Biochem. 2005 Feb;38(2):183-6. doi: 10.1016/j.clinbiochem.2004.10.009.

Authors

Heather A Bentley¹, Dorothy R Belloni, Gregory J Tsongalis

Affiliation

¹ Department of Pathology, Dartmouth Medical School, Dartmouth-Hitchcock Medical Center, One Medical Center Drive, Lebanon, NH 03756, USA.

PMID: 15642283
DOI: 10.1016/j.clinbiochem.2004.10.009

Abstract

Objective: We examined several critical parameters that must be optimized when converting between the ABI Prism 7700 real-time PCR platform and the Cepheid SmartCycler II while using the same primer and probe sequences.

Design and methods: A lyophilized master mix, MgCl(2) concentration, PCR cycling conditions, and ramp times were evaluated.

Results: Optimization of each parameter, including use of the OmniMix HS-lyophilized beads, 6 mM MgCl(2) concentration, changes in PCR cycling parameters, and increased ramp time were necessary to convert this real time PCR assay to a new platform.

Conclusion: We conclude that careful consideration of several analytical parameters can result in a smooth transition of assays between real time PCR platforms.

MeSH terms

Indicators and Reagents
Magnesium Chloride
Polymerase Chain Reaction / instrumentation*
Polymerase Chain Reaction / methods
Polymerase Chain Reaction / standards*
Temperature
Time

Substances

Indicators and Reagents
Magnesium Chloride