Scaling of nucleic acid assays on microelectrophoresis array devices: high-dynamic range multi-gene readout from less than ten transcripts.
In this paper we describe progress in using the prodigious data-collecting ability of multilane microelectrophoresis instruments to bear on problems in scaled nucleic acid assays. We emphasize compound stacking and solid-support loading as means to concentrate <100 pg samples for direct injection. Reaction Mapping is applied to readout quantitative polymerase chain reaction gene-expression and as a way to practically overcome difficulty in interpreting amplification curves of multiplexed quantitative polymerase chain reaction at 20-50 gene/well complexity. We demonstrate multiplexed readout of gene expression over an abundancy range of 9 Log 2 units starting with reverse-transcribed samples as small as five molecules in each sample.
Ueberfeld
Joern
J
Whitehead Institute for Biomedical Research Nine Cambridge Center, Cambridge, MA, USA.
Ehrlich
Daniel J
DJ
D004277:DNA, Single-Stranded / Q000032:analysis*; D046209:Electrophoresis, Microchip / Q000295:instrumentation / Q000379:methods*; D004867:Equipment Design; D020869:Gene Expression Profiling / Q000379:methods*; D055616:Microtechnology; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D004277:DNA, Single-Stranded