Метод фарМакогенетического тестирования после простого выделения днк

Abstract

Background Several commercial DNA isolation kits are available for extracting genomic DNA from whole blood samples, but these procedures require quite some hands-on time and are rather expensive. An alternative technique could be dried blood spot (DBS) sampling, with which DNA isolation is faster, cheaper and logistics are easier. We have developed a non-commercial DBS method and examined whether the quality and quantity of DNA, isolated from DBS and noninvasively obtained buccal swab (BS) samples, was satisfactory. Study design DNA isolation from EDTA blood samples and blood spots on filter paper were compared after DNA isolation with a column method and two different DBS methods. BS samples were obtained by rubbing a sterile, dry cotton swab against the inside of the subject’s cheek. In addition, the quantity of the obtained DNA was measured and melting curve analyses of five cytochrome P450 and three ATP-binding cassette polymorphisms were performed with real-time PCR FRET assays to establish the quality of the obtained DNA from both the DBS and BS samples. Results In all cases the genotype results corresponded completely. Moreover, the derivative melting curves of the DNA samples obtained from the capillary blood and BS were comparable and highly reproducible. The mean DNA concentrations measured were 16.0 ng/μl (12.6-19.4 ng/μl) and 70.2 ng/μl (57.3-83.1 ng/μl), respectively, for DBS and BS samples (p<0.001). Conclusion The DBS DNA isolation appears to be a useful alternative for the commercially available DNA isolation kits and an extremely useful method to discriminate between genotypes. This expands the possibilities of this quick and easy DNA isolation procedure. In particular, the noninvasive BS sampling method appeared to be a good alternative to invasive sampling methods. Simple DNA isolation⏐45