Suspension arrays for high throughput, multiplexed single nucleotide polymorphism genotyping

Citation
B. Armstrong et al., Suspension arrays for high throughput, multiplexed single nucleotide polymorphism genotyping, CYTOMETRY, 40(2), 2000, pp. 102-108
Citations number
20
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Medical Research Diagnosis & Treatment
Journal title
CYTOMETRY
ISSN journal
0196-4763 → ACNP
Volume
40
Issue
2
Year of publication
2000
Pages
102 - 108
Database
ISI
SICI code
0196-4763(20000601)40:2<102:SAFHTM>2.0.ZU;2-W
Abstract
Background: Genetic diversity can help explain disease susceptibility and d ifferential drug response. The most common type of variant is the single nu cleotide polymorphism (SNP). We present a low-cost, high throughput assay f or SNP genotyping. Methods: The assay uses oligonucleotide probes covalently attached to fluor escently encoded microspheres. These probes are hybridized directly to fluo rescently labeled polymerase chain reaction (PCR) products and the results are analyzed in a standard flow cytometer. Results: The genotypes determined with our assay are in good agreement with those determined by TaqMan. The range of G/C content for oligonucleotide p robes mas 23.5-65% in the 17 bases surrounding the SNP. Further optimizatio n of probe length and target concentration is shown to dramatically enhance the assay performance for certain SNPs. Using microspheres which have uniq ue fluorescent signatures, we performed a 32-plex assay where we simultaneo usly determined the genotypes of eight different polymorphic genes. Conclusions: We demonstrate, for the first time, the feasibility of multipl exed genotyping with suspension arrays using direct hybridization analyses. Our approach enables probes to be removed from or added to an array, entra ncing flexibility over conventional chips. The ability to multiplex both th e PCR preparation and the hybridization should enhance the throughput, cost , and speed of the assay. Cytometry 10:102-108, 2000. (C) 2000 Wiley-Liss, Inc.