Collection of buccal cell DNA using treated cards.
We devised a simple, noninvasive, cost-efficient technique for collecting buccal cell DNA for molecular epidemiology studies. Subjects (n = 52) brushed their oral mucosa and expectorated the fluid in their mouths, which was applied to "Guthrie" cards pretreated to retard bacterial growth and inhibit nuclease activity (IsoCode, Schleicher and Schuell, Keene, NH). The cards are well-suited for transport and storage because they dry quickly, need no processing, and are compact and lightweight. We stored the samples at room temperature for 5 days to mimic a field situation and then divided them into portions from which DNA was extracted either immediately or after storage for 9 months at room temperature, -20 degrees C, or -70 degrees C. The fresh samples had a median yield of 2.3 microg of human DNA (range, 0.2-53.8 microg), which was adequate for at least 550 PCR reactions. More than 90% of the samples were amplified in all three beta-globin gene fragment assays attempted. DNA extract frozen for 1 week at -20 degrees C also performed well. Stored samples had reduced DNA yields, which achieved statistical significance for room temperature and -70 degrees C, but not -20 degrees C, storage. However, because all of the stored samples tested were successfully amplified, the observed reduction may represent tighter DNA fixation to the card over time rather than loss of genetic material. We conclude that treated cards are an alternative to brushes/swabs and mouth rinses for the collection of buccal cell DNA and offer some advantages over these methods, particularly for large-scale or large-scale or long-term studies involving stored samples and studies in which samples are collected off-site and transported. Future studies that enable direct comparisons of the various buccal cell collection methods are needed.
Harty
L C
LC
Genetic Epidemiology Branch, National Cancer Institute, Bethesda, Maryland 20892-7236, USA.
Garcia-Closas
M
M
Rothman
N
N
Reid
Y A
YA
Tucker
M A
MA
Hartge
P
P
D002469:Cell Separation / Q000295:instrumentation / Q000379:methods; D004247:DNA / Q000032:analysis*; D005374:Filtration / Q000295:instrumentation; D006801:Humans; D009061:Mouth Mucosa / Q000737:chemistry* / Q000166:cytology*; D009693:Nucleic Acid Hybridization; D010209:Paper; D016133:Polymerase Chain Reaction; D011933:Reagent Kits, Diagnostic
D011933:Reagent Kits, Diagnostic; D004247:DNA