Quality and Quantity of DNA Isolated from Frozen Urine in Population-Based Research

Abstract

In several population-based studies in the past urine samples were collected and stored for future research. We set out to determine the reliability of using such samples for genotyping DNA markers in epidemiologic research. A source of DNA extracted from exfoliated nucleated cells in urine is provided by the DOM cohort, in which specimens were collected 15-25 years ago. We have examined the quality of the DNA in 48 of these samples by measuring the amount of DNA isolated and its ability to provide an adequate PCR template for amplicons of different lengths. MTHFR polymorphism was analyzed in 644 specimens to determine the inter-and intraobserver reproducibility. Although the DNA amount was variable, 26 to 89% of the samples, depending both on the length of the PCR amplicon and on PCR conditions, yielded a visible PCR product. The intra-and interobserver agreements were comparable ( 0.86 and 0.88, respectively). Our results demonstrate that frozen urine samples can be used for DNA typing studies in women after prolonged periods of storage, but with sometimes unpredictable results. Ultimately, the genotype success rate was 89.3%. Urine collection can be considered as a useful method of obtaining DNA in large cohort studies and other circumstances when blood samples cannot be obtained or have not been stored. © 2002 Elsevier Science (USA)

Accuracy of Multiplexed Illumina Platform-Based Single-Nucleotide Polymorphism Genotyping Compared between Genomic and Whole Genome Amplified DNA Collected from Multiple Sources

“…Although we did not evaluate the OmniPlex method for WGA, it is reported to be an alternative that can be used for low-concentration or partially degraded DNA. 4 Indeed, Barker et al (16) found no significant differences in reproducibility or gDNA concordance between the MDA and OmniPlex methods of WGA. It must be noted, however, that their input gDNA was f100 ng and derived from transformed human cell lines (CEPH).…”

“…Additionally, DNA in all samples was extracted from fresh urine; epidemiologic studies are more likely to have archived frozen urine samples. It is unclear what effect the time of day at void, freezing of urine, time stored until freezing, duration of frozen storage, and number of freeze-thaw cycles of stored urine have on subsequent yield and quality of extracted DNA; furthermore, the sex of the urine donor may affect DNA yield (4,6). Urine may be a source of DNA for Illumina genotyping, but we suggest that more data are needed to fully evaluate its use and limitations.…”

“…Recent studies have shown that genomic DNA (gDNA) suitable for SNP genotyping can be obtained from buccal cells and from dried blood spots on Guthrie cards, in addition to well-established sources for gDNA, such as whole blood and buffy coat samples (1,2). gDNA also has been extracted and subsequently genotyped from less concentrated sources, such as urine (3)(4)(5)(6).…”

Cumulative genetic defects in carcinogen metabolism may increase breast cancer risk (The Netherlands)

“…DNA was isolated from urine by alcohol precipitation as described earlier [11]. Urine samples of the study population were thawed overnight at room temperature, mixed vigorously and 50 ml was used for DNA isolation.…”

“…Genotype NAT1 polymorphisms were detected by radioactive dot blot as described previously (11), preceded by PCR using NAT1 specific primers [12]. Briefly, distinction between the NAT1*3, *4, *10 and *11 alleles was carried out by hybridization at 42°C overnight of allele specific oligonucleotides to PCR products spotted on a membrane.…”