Genotyping for DQA1 and PM loci in urine using PCR-based amplification: effects of sample volume, storage temperature, preservatives, and aging on DNA extraction and typing.

摘要: Abstract Urine is often the sample of choice for drug screening in aviation/general forensic toxicology and workplace testing. In some instances, origin submitted samples may be challenged because medicolegal socioeconomic consequences a positive test. Methods individualization biological have reached new boundary with application polymerase chain reaction (PCR) DNA profiling, but successful characterization urine specimens depends on quantity quality present samples. Therefore, study investigated influence storage conditions, volume, concentration modes, extraction procedures, chemical preservations recovered, as well success rate PCR-based genotyping DQA1 PM loci urine. from male female volunteers were divided stored at various temperatures up to 30 days. The results suggested that purification by dialfiltration, using 3000–100,000 molecular weight cut-off filters, did not enhance recovery typing compared simple centrifugation procedures. Extraction urinary organic method resin gave comparable results. Larger volume yielded higher amount DNA, rates affected volumes between 1 5 ml. quantifiable amounts found greater (14–200 ng/ml) than (4–60 decreased elapsed time under both room temperature (RT) frozen storage. Typing also demonstrated RT produced significantly samples, while there was only marginal difference among conditions tested Successful assignment DQA1+PM genotype achieved all fresh urine, independent gender, starting or method. Preservation 0.25% sodium azide acceptable 4°C during period For longer duration, freezing −70°C more appropriate. Thus, applicability clearly