Ultra-rapid DNA analysis using HyBeacon probes and direct PCR amplification from saliva.
We describe a novel probe technology, termed HyBeacons, which provides a new homogeneous method for fluorescence-based sequence detection and allele discrimination. Employing a single nucleotide polymorphism located in the N-acetyltransferase 2 gene as a model system, we demonstrate the utility of HyBeacon probes for rapid and reliable sequence analysis. We also demonstrate that homozygous and heterozygous samples may be accurately identified using a single HyBeacon oligonucleotide. Polymorphic DNA sequences were detected and differentiated by real-time PCR and melt peak methodologies, without performing extraction of genomic DNA prior to target amplification. Employing a combination of homogeneous HyBeacon analysis, the rapid thermal cycling conditions of the LightCycler and direct amplification from saliva, allowed samples to be genotyped within 30 min. Such rapid non-invasive diagnostic technologies may permit 'point-of-care' genetic testing to be performed in hospitals and doctor's surgeries.
French
D J
DJ
BioAnalytical Innovations, LGC (Teddington) Ltd, Queens Road, Teddington, TW11 0LY, UK. dgf@lgc.co.uk
Archard
C L
CL
Andersen
M T
MT
McDowell
D G
DG
D001191:Arylamine N-Acetyltransferase / Q000235:genetics; D004247:DNA / Q000032:analysis*; D004252:DNA Mutational Analysis; D015342:DNA Probes / Q000737:chemistry / Q000592:standards; D005456:Fluorescent Dyes; D005838:Genotype; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D020641:Polymorphism, Single Nucleotide; D012463:Saliva; D013696:Temperature; D013997:Time Factors
D015342:DNA Probes; D005456:Fluorescent Dyes; D004247:DNA; D001191:Arylamine N-Acetyltransferase