Integrated portable polymerase chain reaction-capillary electrophoresis microsystem for rapid forensic short tandem repeat typing.
A portable forensic genetic analysis system consisting of a microfluidic device for amplification and separation of short tandem repeat (STR) fragments as well as an instrument for chip operation and four-color fluorescence detection has been developed. The microdevice performs polymerase chain reaction (PCR) in a 160-nL chamber and capillary electrophoresis (CE) in a 7-cm-long separation channel. The instrumental design integrates PCR thermal cycling, electrophoretic separation, pneumatic valve fluidic control, and four-color laser excited fluorescence detection. A quadruplex Y-chromosome STR typing system consisting of amelogenin and three Y STR loci (DYS390, DYS393, DYS439) was developed and used for validation studies. The multiplex amplification of these 4 loci with 35 PCR cycles followed by CE separation and 4-color fluorescence detection was completed in 1.5 h. All the amplicons can be detected with a limit of detection of 20 copies of male standard DNA in the reactor. Real-world forensic analyses of oral swab and human bone extracts from case evidence were also successfully performed. Mixture analysis demonstrated that a balanced profile can be obtained even at a male-to-female template ratio of 1:10. The successful development and operation of this portable PCR-CE system establishes the feasibility of rapid point-of-analysis DNA typing of forensic casework, of mass disaster samples or of individuals at a security checkpoint.
Liu
Peng
P
UCSF/UC Berkeley Joint Graduate Group in Bioengineering, and Department of Chemistry, University of California, Berkeley, California 94720, USA.
Seo
Tae Seok
TS
Beyor
Nathaniel
N
Shin
Kyoung-Jin
KJ
Scherer
James R
JR
Mathies
Richard A
RA
D041322:Chromosomes, Human, Y / Q000235:genetics; D016172:DNA Fingerprinting / Q000295:instrumentation*; D019075:Electrophoresis, Capillary / Q000295:instrumentation*; D005260:Female; D053803:Forensic Genetics / Q000295:instrumentation*; D006801:Humans; D008297:Male; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation*; D018895:Microsatellite Repeats / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000295:instrumentation*; D012744:Sex Ratio