Proofreading genotyping assays and electrochemical detection of SNPs.
The use of proofreading DNA polymerases in genotyping assays offers the prospect of improved performance. To this end, we have recently used compatible DNA polymerases, protected primers, and substrates to implement proofreading single base extension (P-SBE) and proofreading allele-specific extension (PRASE) assays. Key aspects of the P-SBE and related proofreading assay formats are described here. For transduction of genotyping reactions into physical signals, electrochemical SBE implementations may offer simple, inexpensive assays in electrode array or electrophoretic formats. We have developed electrochemically-labeled nucleotides and electrode detection methods with a view to these applications. Detection of electrochemically-labeled SBE products on an oligonucleotide-modified gold electrode surface is demonstrated.
King
Garry C
GC
School of Biotechnology and Biomolecular Sciences, The University of New South Wales, Sydney, Australia. garry@kinglab.unsw.edu.au
Di Giusto
Daniel A
DA
Wlassoff
Wjatschesslaw A
WA
Giesebrecht
Susanne
S
Flening
Eleanor
E
Tyrelle
Gregory D
GD
D004252:DNA Mutational Analysis / Q000379:methods*; D017931:DNA Primers; D004259:DNA-Directed DNA Polymerase; D004563:Electrochemistry / Q000379:methods*; D004566:Electrodes; D005838:Genotype; D006801:Humans; D009711:Nucleotides / Q000737:chemistry; D010084:Oxidation-Reduction; D020641:Polymorphism, Single Nucleotide
D017931:DNA Primers; D009711:Nucleotides; D004259:DNA-Directed DNA Polymerase