Jornal da Síndrome de Down e Anormalidades Cromossômicas
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Efficient DNA Extraction Protocol for Single Nucleotide Polymorphisms Genotyping in Down Syndrome

Hazel N Barboza, Joissy Aprígio, Carolina Felix Araujo, Marcia G Ribeiro, Marcelo A Costa Lima, Thereza Quirico-Santos and Marcia R Amorim

The collection of buccal epithelial cells derived from saliva has been an alternative source to obtain human DNA, especially from newborns and patients with mental disabilities. Healthy volunteers without history of genetic or neurological disease were included as control to establish adequate conditions for the DNA extraction, storage and genotyping. The study group consisted of 100 Down syndrome (DS) children, 50 with oral motor difficulties. A buccal swab and/or mouthwash procedure before teeth brushing were used for obtaining high-quality genomic DNA for screening gene polymorphisms by conventional and real-time PCR. Comparison of different methods showed that average DNA yield from mouthwash was 343 ng/μl (range 233- 468 ng/μl) and from swabs was 94 ng/μL (range 32-260 ng/μL). Optimized protocol produced high-quality samples that allowed analysis of MTHFR c.677C>T and MTRR c.66A>G polymorphisms by both conventional and Real-time PCR. Extraction of genomic DNA from buccal epithelial cells proved to be a reliable, simple, inexpensive and noninvasive strategy for routine evaluation of genetic variations in DS patients.