Fast multiplexed polymerase chain reaction for conventional and microfluidic short tandem repeat analysis

• Heidi Giese ^[1] ; Roger Lam ^[1] ; Richard Selden ^[1] ; Eugene Tan ^[1]
  1. [1] Network Biosystems
• Localización: Journal of forensic sciences, ISSN-e 1556-4029, ISSN 0022-1198, Vol. 54, Nº. 6, 2009, págs. 1287-1296
• Idioma: inglés
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• Resumen

  • The time required for short tandem repeat (STR) amplification is determined by the temperature ramp rates of the thermal cycler,the components of the reaction mix, and the properties of the reaction vessel. Multiplex amplifications in microfluidic biochip-based and conventionaltube-based thermal cyclers have been demonstrated in 17.3 and 19 min, respectively. Optimized 28-cycle amplification protocols generated alleleswith signal strengths above calling thresholds, heterozygous peak height ratios of greater than 0.65, and incomplete nontemplate nucleotide additionand stutter of less than 15%. Full CODIS-compatible profiles were generated using the Profiler Plus ID, COfiler and Identifiler primer sets. PCR per-formance over a wide range of DNA template levels from 0.006 to 4 ng was characterized by separation and detection on a microfluidic electropho-resis system, Genebench-FXTM. The fast multiplex PCR approach has the potential to reduce process time and cost for STR analysis and enablesdevelopment of a fully integrated microfluidic forensic DNA analysis system