Evaluation of the long-term storage stability of saliva as a source of human DNA
Rent the article at a discountRent now
* Final gross prices may vary according to local VAT.Get Access
The objectives of this paper are to determine the storage stability of saliva at 37 °C over an 18-month period, and its influence on the DNA yield, purity, PCR protocols and genotyping efficacy.
Materials and methods
Of the 60 participants, blood samples were obtained from 10 and saliva from 50. Samples were subjected to different storage conditions: DNA extracted immediately; DNA extracted following storage at 37 °C for 1, 6, 12 and 18 months. Subsequently, DNA yield, OD260/280 and OD260/230 ratios were measured. The isolated DNA was used to amplify exons 0–7 of the RUNX2 gene and subsequently sequenced. Furthermore, 25 SNPs were genotyped.
The mean DNA yield, OD260/280 and OD260/230 ratios obtained from blood were 67.4 ng/μl, 1.8 ± 0.05 and 1.8 ± 0.4 respectively. DNA yield obtained from saliva was significantly higher than blood (p < 0.0001), ranging from 97.4 to 125.8 ng/μl while the OD260/280 ratio ranged from 1.8 ± 0.13 to 1.9 ± 0.1. The success rates for the 25 SNPs ranged from 98 to 100 % for blood and 96–99 % for saliva samples with the genotype frequencies in Hardy–Weinberg equilibrium (>0.01).
Saliva can be stored at 37 °C for 18 months without compromising its quality and ability to endure genetic analyses.
Saliva is a viable source of human DNA to facilitate the feasibility of large-scale genetic studies.
- Ng DP, Koh D, Choo S, Chia KS (2006) Saliva as a viable alternative source of human genomic DNA in genetic epidemiology. Clin Chim Acta 367:81–85 CrossRef
- Ng DP, Koh D, Choo S, Ng V, Fu QY (2004) Effect of storage conditions on the extraction of PCR-quality genomic DNA from saliva. Clin Chim Acta 343:191–194 CrossRef
- Quinque D, Kittler R, Kayser M, Stoneking M, Nasidze I (2006) Evaluation of saliva as a source of human DNA for population and association studies. Anal Biochem 353:272–277 CrossRef
- Rogers NL, Cole SA, Lan HC, Crossa A, Demerath EW (2007) New saliva DNA collection method compared to buccal cell collection techniques for epidemiological studies. Am J Hum Biol 19:319–326 CrossRef
- Mulot C, Stücker I, Clavel J, Beaune P, Loriot MA (2005) Collection of human genomic DNA from buccal cells for genetics studies: comparison between cytobrush, mouthwash, and treated card. J Biomed Biotechnol 3:291–296 CrossRef
- García-Closas M, Egan KM, Abruzzo J, Newcomb PA, Titus-Ernstoff L, Franklin T et al (2001) Collection of genomic DNA from adults in epidemiological studies by buccal cytobrush and mouthwash. Cancer Epidemiol Biomarkers Prev 6:687–696
- Cozier YC, Palmer JR, Rosenberg L (2004) Comparison of methods for collection of DNA samples by mail in the Black Women’s Health Study. Ann Epidemiol 2:117–122 CrossRef
- Luan JA, Wong MY, Day NE, Wareham NJ (2001) Sample size determination for studies of gene–environment interaction. Int J Epidemiol 30:1035–1040 CrossRef
- Holland NT, Smith MT, Eskenazi B, Bastaki M (2003) Biological sample collection and processing for molecular epidemiological studies. Mutat Res 543:217–234 CrossRef
- Xuan D, Li S, Zhang X, Hu F, Lin L, Wang C et al (2008) Mutations in the RUNX2 gene in Chinese patients with cleidocranial dysplasia. Ann Clin Lab Sci 38:15–24
- Phillips MS, Lawrence R, Sachidanandam R, Morris AP, Balding DJ, Donaldson MA et al (2003) Chromosome-wide distribution of haplotype blocks and the role of recombination hot spots. Nat Genet 33:382–387 CrossRef
- Halsall A, Ravetto P, Reyes Y, Thelwell N, Davidson A, Gaut R et al (2008) The quality of DNA extracted from liquid or dried blood is not adversely affected by storage at 4 degrees C for up to 24 h. Int J Epidemiol 37(Suppl 1):i7–i10 CrossRef
- Visvikis S, Schlenck A, Maurice M (1998) DNA extraction and stability of epidemiological studies. Clin Chem Lab Med 36:551–555 CrossRef
- Dawes C (2003) Estimates, from salivary analyses, of the turnover time of the oral mucosal epithelium in humans and the number of bacteria in an edentulous mouth. Arch Oral Biol 48:329–336 CrossRef
- Halsall A, Ravetto P, Reyes Y, Thelwell N, Davidson A, Gaut R et al (2008) The quality of DNA extracted from liquid or dried blood is not adversely affected by storage at 4 degrees C for up to 24 h. Int J Epidemiol 37:7–10 CrossRef
- Rylander-Rudqvist T, Håkansson N, Tybring G, Wolk A (2006) Quality and quantity of saliva DNA obtained from the self-administrated oragene method—a pilot study on the cohort of Swedish men. Cancer Epidemiol Biomarkers Prev 15:1742–1745 CrossRef
- Chartier J, Birnboim H (2004) Bacterial DNA content with OrageneTM. DNA Genotek, Ottawa
- van Schie RC, Wilson ME (1997) Saliva: a convenient source of DNA for analysis of bi-allelic polymorphisms of Fc gamma receptor IIA (CD32) and Fc gamma receptor IIIB (CD16). J Immunol Methods 208:91–101 CrossRef
- Andrisin TE, Humma LM, Johnson JA (2002) Collection of genomic DNA by the non-invasive mouthwash method for use in pharmacogenetic studies. Pharmacotherapy 22:954–960 CrossRef
- Evaluation of the long-term storage stability of saliva as a source of human DNA
Clinical Oral Investigations
Volume 17, Issue 7 , pp 1719-1725
- Cover Date
- Print ISSN
- Online ISSN
- Springer Berlin Heidelberg
- Additional Links
- Industry Sectors
- Author Affiliations
- 1. Paediatric Dentistry, School of Dentistry, The University of Western Australia, Perth, Australia
- 3. Paediatric Dentistry, School of Dentistry, Oral Health Centre of Western Australia, 17 Monash Avenue, Nedlands, WA, 6009, Australia
- 2. Paediatric Dentistry and Orthodontics, Faculty of Dentistry, The University of Hong Kong, Hong Kong, SAR, China