Buccal swabs and treated cards: methodological considerations for molecular epidemiologic studies examining pediatric populations.
Self-collection of buccal cells provides a noninvasive method for obtaining biologic samples for genetic analyses in pediatric studies. Nevertheless, low yields, microbial contamination, and degradation of buccal samples present challenges for epidemiologic studies incorporating genetic investigations. The aims of this study were to compare the quality and yield of DNA extracted from buccal specimens with BuccalAmp swabs (Epicenter BioTechnologies, Madison, Wisconsin) or FTA cards (Whatman, Inc., Clifton, New Jersey) and to investigate the use of whole-genome amplification (WGA) for increasing DNA yields for single nucleotide polymorphism analyses. Buccal specimens were collected from 55 children with acute lymphoblastic leukemia and 52 control children without acute lymphoblastic leukemia in New South Wales, Australia, in 2003-2004. Real-time polymerase chain reaction was used to evaluate polymorphisms in the genes encoding the cytochrome p450 enzyme CYP3A4 (CYP3A4 A392G, also known as CYP3A4*1B) and the steroid xenobiotic receptor (SXR C25385T). Results showed that DNA could be isolated from buccal specimens collected by use of both methods and that yields could be substantially improved with WGA without introducing genotyping error. However, DNA quality was poorer in samples collected by BuccalAmp swabs, and the presence of polymerase chain reaction inhibitors in these samples reduced the sensitivity of quantitative real-time PCR analysis. These findings show that different methods for collecting buccal samples impact on the downstream success of genetic investigations and influence DNA quality after WGA.
Beckett
Sara M
SM
Children's Cancer Institute Australia for Medical Research, Randwick, New South Wales, Australia.
Laughton
Stephen J
SJ
Pozza
Luciano Dalla
LD
McCowage
Geoffrey B
GB
Marshall
Glenn
G
Cohn
Richard J
RJ
Milne
Elizabeth
E
Ashton
Lesley J
LJ
D002610:Cheek; D002648:Child; D004247:DNA / Q000032:analysis*; D005260:Female; D005838:Genotype; D006801:Humans; D008297:Male; D017720:Molecular Epidemiology / Q000379:methods*; D009061:Mouth Mucosa / Q000166:cytology*; D021141:Nucleic Acid Amplification Techniques / Q000295:instrumentation / Q000379:methods*; D010209:Paper; D010372:Pediatrics / Q000379:methods; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide; D017422:Sequence Analysis, DNA / Q000379:methods*; D013048:Specimen Handling / Q000379:methods
D004247:DNA