Telomere length measurement on the Roche LightCycler 480 platform

Abstract

Background: The average length of telomeres as measured in genomic DNA from human peripheral blood leukocytes is proving to be a potential biomarker of great interest, particularly with respect to studies of aging, specific diseases, and the effects of various stresses on overall health. Aims: The aim of this study was to establish an effective real-time quantitative polymerase chain reaction (qPCR) method for telomere length measurement on the Roche LightCycler® 480 (LC480) real-time PCR platform. Methods: Measurement of relative average telomere length was achieved by comparing products amplified from telomere-specific primers and single copy reference gene primers in a ratio (T/S). Results: Extensive testing led us to conclude that a modification of the original two-plate T/S assay was more compatible with this platform than the recently developed single-plate assay, and that choice of hot-start Taq polymerase and intercalating dye were critical factors. Conclusions: This modified assay generates reliable measurements as judged by correlation with data derived by the telomeric restriction fragment Southern blot-based method.