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Choice of gDNA isolation method has a significant impact on average murine Telomere Length estimates.

Kidd, E; Meimaridou, E; Williams, J; Metherell, LA; Walley, AJ; Fairbrother, UL (2024) Choice of gDNA isolation method has a significant impact on average murine Telomere Length estimates. Prep Biochem Biotechnol, 54 (6). pp. 788-795. ISSN 1532-2297 https://doi.org/10.1080/10826068.2023.2288572
SGUL Authors: Walley, Andrew John

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Abstract

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 to 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 a T/S ratio. We use a monochromatic-multiplex-qPCR (mmqPCR) method which multiplexes the PCR and enables quantification of the target and the single copy gene within the same qPCR reaction. We demonstrate that 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. 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.

Item Type: Article
Additional Information: © 2023 The Author(s). Published with license by Taylor & Francis Group, LLC This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The terms on which this article has been published allow the posting of the Accepted Manuscript in a repository by the author(s) or with their consent.
Keywords: DNA isolation, magnetic beads, mmqPCR, phenol/chloroform, spin columns, telomere length, 03 Chemical Sciences, 06 Biological Sciences, 10 Technology, Biotechnology
SGUL Research Institute / Research Centre: Academic Structure > Institute of Medical & Biomedical Education (IMBE)
Academic Structure > Institute of Medical & Biomedical Education (IMBE) > Centre for Biomedical Education (INMEBE)
Journal or Publication Title: Prep Biochem Biotechnol
ISSN: 1532-2297
Language: eng
Dates:
DateEvent
1 July 2024Published
13 December 2023Published Online
Publisher License: Creative Commons: Attribution 4.0
PubMed ID: 38088914
Go to PubMed abstract
URI: https://openaccess.sgul.ac.uk/id/eprint/115937
Publisher's version: https://doi.org/10.1080/10826068.2023.2288572

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