Choice of Isolation method has a significant impact on average murine Telomere Length estimates

Abstract

Abstract Background Telomere Length (TL) and integrity is significantly associated with age-related disease, multiple genetic and environmental factors. We observe mouse genomic DNA (gDNA) isolation methods have a significant impact on average TL estimates. The canonical qPCR method does not measure TL directly but via the ratio of telomere repeats to a single copy gene (SCG) generating an TS ratio. We use an mmqPCR method which multiplexes the PCR and enables quantification of the target and the single copy gene within the same qPCR reaction. Results We demonstrate TL measurements, from murine gDNA, isolated via Spin Columns (SC) and Magnetic Beads (MB), generate significantly smaller T/S ratios compared to gDNA isolated via traditional phenol/chloroform methods. The former methods may impede correct TL estimation by producing non representative fragment sets and reducing qPCR efficacy. Conclusions This work highlights discrepancies in TL measurements due to different extraction techniques. We recommend the use of gDNA isolation methods that are shown to preserve DNA length and integrity, such as phenol/chloroform isolation. We propose that widely used high throughput DNA isolation methodologies can create spurious associations within a sample set, thus creating misleading data. We suggest that published TL associations should be revisited in the light of these data.