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Cancer Epidemiology, Biomarkers & Prevention
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Letters to the Editor

Telomere Length Varies by DNA Extraction Method: Implications for Epidemiologic Research—Letter

Jonathan N. Hofmann, Amy A. Hutchinson, Richard Cawthon, Chin-San Liu, Shannon M. Lynch, Qing Lan, Nathaniel Rothman, Rachael Stolzenberg-Solomon and Mark P. Purdue
Jonathan N. Hofmann
1Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda; 2Cancer Genomics Research Laboratory, NCI-DCEG, Leidos Biomedical Research, Inc., Frederick, Maryland; 3Department of Human Genetics, University of Utah, Salt Lake City, Utah; 4Center for Clinical Epidemiology and Biostatistics, Center for Genetics and Complex Traits, University of Pennsylvania, Philadelphia, Pennsylvania; and 5Vascular and Genomic Research Center, Changhua Christian Hospital, Changhua, Taiwan
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Amy A. Hutchinson
1Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda; 2Cancer Genomics Research Laboratory, NCI-DCEG, Leidos Biomedical Research, Inc., Frederick, Maryland; 3Department of Human Genetics, University of Utah, Salt Lake City, Utah; 4Center for Clinical Epidemiology and Biostatistics, Center for Genetics and Complex Traits, University of Pennsylvania, Philadelphia, Pennsylvania; and 5Vascular and Genomic Research Center, Changhua Christian Hospital, Changhua, Taiwan
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Richard Cawthon
1Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda; 2Cancer Genomics Research Laboratory, NCI-DCEG, Leidos Biomedical Research, Inc., Frederick, Maryland; 3Department of Human Genetics, University of Utah, Salt Lake City, Utah; 4Center for Clinical Epidemiology and Biostatistics, Center for Genetics and Complex Traits, University of Pennsylvania, Philadelphia, Pennsylvania; and 5Vascular and Genomic Research Center, Changhua Christian Hospital, Changhua, Taiwan
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Chin-San Liu
1Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda; 2Cancer Genomics Research Laboratory, NCI-DCEG, Leidos Biomedical Research, Inc., Frederick, Maryland; 3Department of Human Genetics, University of Utah, Salt Lake City, Utah; 4Center for Clinical Epidemiology and Biostatistics, Center for Genetics and Complex Traits, University of Pennsylvania, Philadelphia, Pennsylvania; and 5Vascular and Genomic Research Center, Changhua Christian Hospital, Changhua, Taiwan
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Shannon M. Lynch
1Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda; 2Cancer Genomics Research Laboratory, NCI-DCEG, Leidos Biomedical Research, Inc., Frederick, Maryland; 3Department of Human Genetics, University of Utah, Salt Lake City, Utah; 4Center for Clinical Epidemiology and Biostatistics, Center for Genetics and Complex Traits, University of Pennsylvania, Philadelphia, Pennsylvania; and 5Vascular and Genomic Research Center, Changhua Christian Hospital, Changhua, Taiwan
1Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda; 2Cancer Genomics Research Laboratory, NCI-DCEG, Leidos Biomedical Research, Inc., Frederick, Maryland; 3Department of Human Genetics, University of Utah, Salt Lake City, Utah; 4Center for Clinical Epidemiology and Biostatistics, Center for Genetics and Complex Traits, University of Pennsylvania, Philadelphia, Pennsylvania; and 5Vascular and Genomic Research Center, Changhua Christian Hospital, Changhua, Taiwan
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Qing Lan
1Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda; 2Cancer Genomics Research Laboratory, NCI-DCEG, Leidos Biomedical Research, Inc., Frederick, Maryland; 3Department of Human Genetics, University of Utah, Salt Lake City, Utah; 4Center for Clinical Epidemiology and Biostatistics, Center for Genetics and Complex Traits, University of Pennsylvania, Philadelphia, Pennsylvania; and 5Vascular and Genomic Research Center, Changhua Christian Hospital, Changhua, Taiwan
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Nathaniel Rothman
1Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda; 2Cancer Genomics Research Laboratory, NCI-DCEG, Leidos Biomedical Research, Inc., Frederick, Maryland; 3Department of Human Genetics, University of Utah, Salt Lake City, Utah; 4Center for Clinical Epidemiology and Biostatistics, Center for Genetics and Complex Traits, University of Pennsylvania, Philadelphia, Pennsylvania; and 5Vascular and Genomic Research Center, Changhua Christian Hospital, Changhua, Taiwan
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Rachael Stolzenberg-Solomon
1Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda; 2Cancer Genomics Research Laboratory, NCI-DCEG, Leidos Biomedical Research, Inc., Frederick, Maryland; 3Department of Human Genetics, University of Utah, Salt Lake City, Utah; 4Center for Clinical Epidemiology and Biostatistics, Center for Genetics and Complex Traits, University of Pennsylvania, Philadelphia, Pennsylvania; and 5Vascular and Genomic Research Center, Changhua Christian Hospital, Changhua, Taiwan
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Mark P. Purdue
1Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda; 2Cancer Genomics Research Laboratory, NCI-DCEG, Leidos Biomedical Research, Inc., Frederick, Maryland; 3Department of Human Genetics, University of Utah, Salt Lake City, Utah; 4Center for Clinical Epidemiology and Biostatistics, Center for Genetics and Complex Traits, University of Pennsylvania, Philadelphia, Pennsylvania; and 5Vascular and Genomic Research Center, Changhua Christian Hospital, Changhua, Taiwan
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DOI: 10.1158/1055-9965.EPI-14-0145 Published June 2014
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In their recent article, Cunningham and colleagues (1) reported differences in leukocyte telomere length (TL) related to the method of DNA extraction, with shorter TL measurements among samples extracted using QIAamp (Qiagen) compared with those extracted using PureGene or phenol/chloroform methods. It is unclear whether such within-subject differences are also observed with other commonly used methods of DNA extraction, such as the Promega ReliaPrep Kit, or for other suspected DNA-based biomarkers of cancer risk, such as mitochondrial DNA (mtDNA) copy number.

To address these questions, we conducted a similar methodologic evaluation involving paired samples of genomic DNA freshly extracted from the same buffy coat source specimens using two different methods: the QIAamp DNA Blood Midi Kit from Qiagen and the ReliaPrep Large Volume HT gDNA Isolation Kit from Promega. The QIAamp Kit uses a standard column matrix for DNA capture and elution, while the ReliaPrep chemistry is based on magnetic bead capture of nucleic acid. We measured leukocyte TL in paired samples from 40 subjects and mtDNA copy number in paired samples from 48 subjects in the Research Donor Program at the Frederick National Laboratory for Cancer Research (Frederick, MD). TL and mtDNA copy number were measured in triplicate relative to nuclear DNA using quantitative PCR (qPCR); assay methods have been described previously (2, 3). Masked replicate QC samples (N = 8) from a single subject were interspersed to assess assay reproducibility; coefficients of variation were very low and did not differ by extraction method (TL: 5.4% for QIAamp and 5.1% for ReliaPrep; mtDNA copy number: 3.8% for QIAamp and 4.4% for ReliaPrep).

As shown in Table 1, we found that samples extracted using QIAamp had significantly shorter leukocyte TL compared with those extracted using ReliaPrep (medians of 1.13 and 1.48, respectively; P < 0.001). Conversely, for mtDNA copy number, levels were significantly higher in samples extracted using QIAamp compared with ReliaPrep (medians of 212 and 184, respectively; P = 0.005). The correlation between paired samples was moderately high for TL (Spearman ρ = 0.71) and weaker for mtDNA copy number (Spearman ρ = 0.46).

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Table 1.

Differences in leukocyte TL and mtDNA copy number by DNA extraction method in paired samples from the same subjects

Our data corroborate the findings of Cunningham and colleagues and underscore the importance of taking DNA extraction method into consideration in epidemiologic studies investigating TL or mtDNA copy number in relation to cancer and other chronic diseases. Whenever possible, all the samples in a given study should be extracted using the same method to ensure comparability between subjects in the measurements of these analytes.

See the Response, p. 1131

Disclosure of Potential Conflicts of Interest

R. Cawthon has ownership interest (including patents) in a patent on the qPCR method of measuring average TL and is a consultant/advisory board member of Telomere Diagnostics, Inc. No potential conflicts of interest were disclosed by the other authors.

  • Received February 6, 2014.
  • Accepted February 12, 2014.
  • ©2014 American Association for Cancer Research.

References

  1. 1.↵
    1. Cunningham JM,
    2. Johnson RA,
    3. Litzelman K,
    4. Skinner HG,
    5. Seo S,
    6. Engelman CD,
    7. et al.
    Telomere length varies by DNA extraction method: implications for epidemiologic research. Cancer Epidemiol Biomarkers Prev 2013;22:2047–54.
    OpenUrlAbstract/FREE Full Text
  2. 2.↵
    1. Cawthon RM
    . Telomere length measurement by a novel monochrome multiplex quantitative PCR method. Nucleic Acids Res 2009;37:e21.
    OpenUrlAbstract/FREE Full Text
  3. 3.↵
    1. Liu CS,
    2. Tsai CS,
    3. Kuo CL,
    4. Chen HW,
    5. Lii CK,
    6. Ma YS,
    7. et al.
    Oxidative stress-related alteration of the copy number of mitochondrial DNA in human leukocytes. Free Radic Res 2003;37:1307–17.
    OpenUrlCrossRefPubMed
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Cancer Epidemiology Biomarkers & Prevention: 23 (6)
June 2014
Volume 23, Issue 6
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Telomere Length Varies by DNA Extraction Method: Implications for Epidemiologic Research—Letter
Jonathan N. Hofmann, Amy A. Hutchinson, Richard Cawthon, Chin-San Liu, Shannon M. Lynch, Qing Lan, Nathaniel Rothman, Rachael Stolzenberg-Solomon and Mark P. Purdue
Cancer Epidemiol Biomarkers Prev June 1 2014 (23) (6) 1129-1130; DOI: 10.1158/1055-9965.EPI-14-0145

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Telomere Length Varies by DNA Extraction Method: Implications for Epidemiologic Research—Letter
Jonathan N. Hofmann, Amy A. Hutchinson, Richard Cawthon, Chin-San Liu, Shannon M. Lynch, Qing Lan, Nathaniel Rothman, Rachael Stolzenberg-Solomon and Mark P. Purdue
Cancer Epidemiol Biomarkers Prev June 1 2014 (23) (6) 1129-1130; DOI: 10.1158/1055-9965.EPI-14-0145
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