Telomere Length Varies By DNA Extraction Method: Implications for Epidemiologic Research

Abstract

Background Both shorter and longer telomeres in peripheral blood leukocyte (PBL) DNA have been associated with cancer risk. However, associations remain inconsistent across studies of the same cancer type. This study compares DNA preparation methods to determine telomere length from colorectal cancer patients. Methods We examined PBL relative telomere length (RTL) measured by quantitative PCR (qPCR) in 1,033 colorectal cancer patients and 2,952 healthy controls. DNA was extracted with Phenol/Chloroform, PureGene or QIAamp. Results We observed differences in RTL depending on DNA extraction method (p<0.001). Phenol/Chloroform extracted DNA had a mean RTL (T/S ratio) of 0.78 (range 0.01-6.54) ) compared to PureGene extracted DNA (mean RTL of 0.75; range 0.00-12.33). DNA extracted by QIAamp yielded a mean RTL of 0.38 (range 0.02-3.69). We subsequently compared RTL measured by qPCR from an independent set of 20 colorectal cancer cases and 24 normal controls in PBL DNA extracted by each of the three extraction methods. The range of RTL measured by qPCR from QIAamp-extracted DNA (0.17-0.58-) was smaller than from either PureGene or Phenol/Chloroform (ranges:0.04-2.67 and 0.32-2.81, respectively). Conclusions RTL measured by qPCR from QIAamp-extracted DNA was smaller than from either PureGene or Phenol/Chloroform (p<0.001). Impact Differences in DNA extraction method may contribute to the discrepancies between studies seeking to find an association between the risk of cancer or other diseases and RTL.

Genetic Variants Related to Longer Telomere Length are Associated with Increased Risk of Renal Cell Carcinoma

“…All studies measured telomere length from a single time point, which may not adequately reflect telomere length status in the etiologically relevant time window, and were susceptible to confounding from RCC risk factors that may be associated with telomere length such as smoking [13,18] and obesity [19]. Furthermore, qPCR-based measurements of telomere length are sensitive to preanalytic factors such as DNA source material and extraction method [12,20,21]. …”

Psychiatric disorders and leukocyte telomere length: Underlying mechanisms linking mental illness with cellular aging

“…Important caveats must be considered in interpreting LTL (Aviv et al, 2006; Epel, 2012), e.g., (a) distinguishing between telomere shortening in the individual leukocyte vs. “apparent” telomere shortening when examining average LTL, due to a re-distribution of leukocyte cell types having different telomere lengths, such as naïve vs. memory T cells; (b) the relationship of LTL to TL in other tissues has not been well established, and TL varies by tissue (although TL is generally positively correlated across certain, but not all, tissues within individuals) (Daniali et al, 2013; Dlouha et al, 2014; Friedrich et al, 2000; Gadalla et al, 2010; Lukens et al, 2009; Mitchell et al, 2014; Nakamura et al, 2002; Takubo et al, 2002; Takubo et al, 2010; Thomas et al, 2008); (c) different results may derive from different DNA extraction and assay methods and different laboratories (Aubert et al, 2012; Aviv et al, 2011; Cunningham et al, 2013; Montpetit et al, 2014; Nieratschker et al, 2013); and (d) even slight DNA degradation can yield spurious TL measuements (Dlouha et al, 2014). Further, many subject-level variables, such as age, sex, genetic polymorphisms, “resiliency,” education, history of early life adversities, parental responsiveness, socioeconomic status, health behaviors, diet, and latent or active viral infections (e.g., cytomegalovirus or herpes virus) may affect LTL, independent of the disease process being studied (Adler et al, 2013; Asok et al, 2013; Aviv et al, 2011; Effros, 2011; Eitan et al, 2014; Epel, 2012; Epel, 2009; Gardner et al, 2014; Gutierrez-Rodrigues et al, 2014; Jacobs et al, 2013; Lung et al, 2005; Nieratschker et al, 2013; Price et al, 2012; Puterman et al, 2013; Puterman et al, 2010; Rizzo et al, 2013; Spyridopoulos et al, 2009; Wikgren et al, 2012a).…”

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

“…In their recent article (1), Cunningham and colleagues 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 to 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.…”