Degenerate oligonucleotide primed-polymerase chain reaction and capillary electrophoretic analysis of human DNA on microchip-based devices.
Random amplification of the human genome using the degenerate oligonucleotide primed-polymerase chain reaction (DOP-PCR) was performed in a silicon-glass chip. An aliquot of the DOP-PCR amplified genomic DNA was then introduced into another silicon-glass chip for a locus-specific, multiplex PCR of the dystrophin gene exons in order to detect deletions causing Duchenne/Becker muscular dystrophy. Amplicons were analyzed by both conventional capillary electrophoresis and microchip electrophoresis and results were compared to those obtained using standard non-chip-based PCR assays. Results from microchip electrophoresis were consistent with those from conventional capillary electrophoresis. Whole genome amplification products obtained by DOP-PCR proved to be a suitable template for multiplex PCR as long as amplicon size was < 250 bp. Successful detection and resolution of all PCR products from the multiplex PCR clearly shows the feasibility of performing complex PCR assays using microfabricated devices.
Cheng
J
J
Department of Pathology, University of Pennsylvania, Philadelphia, USA.
Waters
L C
LC
Fortina
P
P
Hvichia
G
G
Jacobson
S C
SC
Ramsey
J M
JM
Kricka
L J
LJ
Wilding
P
P
D004247:DNA / Q000032:analysis* / Q000235:genetics; D017931:DNA Primers; D016189:Dystrophin / Q000235:genetics; D004587:Electrophoresis, Agar Gel; D019075:Electrophoresis, Capillary / Q000295:instrumentation; D015894:Genome, Human; D005898:Glass; D006801:Humans; D008297:Male; D008851:Micropore Filters; D009136:Muscular Dystrophies / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000295:instrumentation / Q000379:methods; D017384:Sequence Deletion; D012825:Silicon
D017931:DNA Primers; D016189:Dystrophin; D004247:DNA; D012825:Silicon