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Fast Multiplexed Polymerase Chain Reaction for Conventional and Microfluidic Short Tandem Repeat Analysis
Author(s) -
Giese Heidi,
Lam Roger,
Selden Richard,
Tan Eugene
Publication year - 2009
Publication title -
journal of forensic sciences
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.715
H-Index - 96
eISSN - 1556-4029
pISSN - 0022-1198
DOI - 10.1111/j.1556-4029.2009.01200.x
Subject(s) - str analysis , microfluidics , multiplex , microsatellite , multiplex polymerase chain reaction , polymerase chain reaction , biochip , primer (cosmetics) , tandem , analytical chemistry (journal) , materials science , microbiology and biotechnology , chromatography , chemistry , biology , genetics , nanotechnology , allele , gene , organic chemistry , composite material
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 conventional tube‐based thermal cyclers have been demonstrated in 17.3 and 19 min, respectively. Optimized 28‐cycle amplification protocols generated alleles with signal strengths above calling thresholds, heterozygous peak height ratios of greater than 0.65, and incomplete nontemplate nucleotide addition and stutter of less than 15%. Full CODIS‐compatible profiles were generated using the Profiler Plus ID, COfiler and Identifiler primer sets. PCR performance over a wide range of DNA template levels from 0.006 to 4 ng was characterized by separation and detection on a microfluidic electrophoresis system, Genebench‐FX ™ . The fast multiplex PCR approach has the potential to reduce process time and cost for STR analysis and enables development of a fully integrated microfluidic forensic DNA analysis system.