title: Integrated DNA purification, PCR, sample cleanup, and capillary electrophoresis microchip for forensic human identification.
abstract: A fully integrated microdevice and process for forensic short tandem repeat (STR) analysis has been developed that includes sequence-specific DNA template purification, polymerase chain reaction (PCR), post-PCR cleanup and inline injection, and capillary electrophoresis (CE). Fragmented genomic DNA is hybridized with biotin-labeled capture oligos and pumped through a fluidized bed of magnetically immobilized streptavidin-coated beads in microchannels where the target DNA is bound to the beads. The bead-DNA conjugates are then transferred into a 250 nL PCR reactor for autosomal STR amplification using one biotin and one fluorescence-labeled primer. The resulting biotin-labeled PCR products are electrophoretically injected through a streptavidin-modified capture gel where they are captured to form a concentrated and purified injection plug. The thermally released sample plug is injected into a 14 cm long CE column for fragment separation and detection. The DNA template capture efficiency provided by the on-chip sequence-specific template purification is determined to be 5.4% using K562 standard DNA. This system can produce full 9-plex STR profiles from 2.5 ng input standard DNA and obtain STR profiles from oral swabs in about 3 hours. This fully integrated microsystem with sample-in-answer-out capability is a significant advance in the development of rapid, sensitive, and reliable micro-total analysis systems for on-site human identification.
authors.affiliation: UCSF/UC Berkeley Joint Graduate Group in Bioengineering, University of California, Berkeley, CA 94720, USA.
authors.forename: Peng
authors.initials: P
authors.lastname: Liu
authors.forename: Xiujun
authors.initials: X
authors.lastname: Li
authors.forename: Susan A
authors.initials: SA
authors.lastname: Greenspoon
authors.forename: James R
authors.initials: JR
authors.lastname: Scherer
authors.forename: Richard A
authors.initials: RA
authors.lastname: Mathies
mesh_terms: D004247:DNA / Q000737:chemistry* / Q000302:isolation & purification; D046209:Electrophoresis, Microchip / Q000295:instrumentation / Q000379:methods*; D053803:Forensic Genetics / Q000379:methods*; D006801:Humans; D046208:Microchip Analytical Procedures; D018895:Microsatellite Repeats; D016133:Polymerase Chain Reaction
chemical_list: D004247:DNA