silicon and Muscular-Dystrophies

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.

Topics: DNA; DNA Primers; Dystrophin; Electrophoresis, Agar Gel; Electrophoresis, Capillary; Genome, Human; Glass; Humans; Male; Micropore Filters; Muscular Dystrophies; Polymerase Chain Reaction; Sequence Deletion; Silicon

Microfabricated devices containing arrays of nucleic acid hybridization sites, known as genosensors, are being developed for a variety of uses in genomic analysis. A great deal of the overall genosensor development effort involves optimization of experimental conditions in the actual use of genosensors. Here we describe a "low-tech" form of genosensor technology, involving arrays of oligonucleotides on glass microscope slides, which can be used to define optimal operating conditions and to develop applications of hybridization arrays in genome mapping and sequencing. In addition, we describe a porous silicon genosensor, which can be operated in a flowthrough mode, and discuss its advantages over current flat-surface designs. Porous silicon genosensors containing arrays of DNA fragments offer several unique capabilities in genome analysis.

Topics: Base Sequence; Biosensing Techniques; Chromosome Mapping; DNA; Glass; Hot Temperature; Humans; Molecular Sequence Data; Muscular Dystrophies; Nucleic Acid Hybridization; Oligonucleotide Probes; Polymerase Chain Reaction; Sequence Analysis, DNA; Silicon