Determinants of DNA yield and quality from buccal cell samples collected with mouthwash.
Buccal cells are becoming an important source of genomic DNA in epidemiological studies, but little is known about the effect of different sampling conditions on DNA quality and yield. We used a mouthwash protocol to collect six daily buccal cell samples from 35 healthy volunteers. Twenty-four individuals (six men and 18 women) correctly completed the protocol and were included in paired analyses to determine whether "swish" time (30 s versus 60 s), toothbrushing before collection, or lag time between collection and DNA extraction (1 day versus 5, 10, or 30 days at room temperature) would affect sample quality and yield. Total DNA, human-specific DNA (hDNA), degradation of DNA, and ability to amplify by PCR were determined. hDNA yield did not significantly vary by "swish" time. However, toothbrushing 1 h before sample collection reduced the amount of hDNA by nearly 40% (34 microg versus 21 microg; P = 0.06). Median hDNA yields for samples that were held for 1, 5, 10, and 30 days before extraction were 32 microg (range, 4-196), 32 microg (2-194), 23 microg (3-80), and 21 microg (5-56), respectively. The 10- and 30-day samples had significantly less hDNA than those processed after 1 day (P = 0.01). PCR success rates for beta-globin gene fragments of length 268 bp, 536 bp, and 989 bp were 94% or better, and high molecular weight DNA (>23 kb) was found in all but one sample. These results suggest that buccal cells should be collected before brushing teeth and processed within 5 days of collection to maximize hDNA yield.
Feigelson
H S
HS
Department of Epidemiology and Surveillance Research, American Cancer Society, National Home Office, Atlanta, Georgia 30329-4251, USA. heather.feigelson@cancer.org
Rodriguez
C
C
Robertson
A S
AS
Jacobs
E J
EJ
Calle
E E
EE
Reid
Y A
YA
Thun
M J
MJ
D000328:Adult; D004247:DNA / Q000032:analysis / Q000302:isolation & purification*; D016021:Epidemiologic Studies; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D009061:Mouth Mucosa / Q000166:cytology*; D009067:Mouthwashes; D010865:Pilot Projects; D016133:Polymerase Chain Reaction; D012016:Reference Values; D015203:Reproducibility of Results; D013048:Specimen Handling
D009067:Mouthwashes; D004247:DNA