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A Microsphere-Based Assay for Multiplexed Single Nucleotide Polymorphism Analysis Using Single Base Chain Extension
Open accessAbstract
A rapid, high throughput readout for single-nucleotide polymorphism (SNP) analysis was developed employing single base chain extension and cytometric analysis of an array of fluorescent microspheres. An array of fluorescent microspheres was coupled with uniquely identifying sequences, termed complementary ZipCodes (cZipCodes), which allowed for multiplexing possibilities. For a given assay, querying a polymorphic base involved extending an oligonucleotide containing both a ZipCode and a SNP-specific sequence with a DNA polymerase and a pair of fluoresceinated dideoxynucleotides. To capture the reaction products for analysis, the ZipCode portion of the oligonucleotide was hybridized with its cZipCodes on the microsphere. Flow cytometry was used for microsphere decoding and SNP typing by detecting the fluorescein label captured on the microspheres. In addition to multiplexing capability, the ZipCode system allows multiple sets of SNPs to be analyzed by a limited set of cZipCode-attached microspheres. A standard set of non-cross reactive ZipCodes was established experimentally and the accuracy of the system was validated by comparison with genotypes determined by other technologies. From a total of 58 SNPs, 55 SNPs were successfully analyzed in the first pass using this assay format and all 181 genotypes across the 55 SNPs were correct. These data demonstrate that the microsphere-based single base chain extension (SBCE) method is a sensitive and reliable assay. It can be readily adapted to an automated, high-throughput genotyping system. [Primer sequences used in this study are available as online supplementary materials at www.genome.org.]
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Association of apolipoprotein E allele ϵ4 with late‐onset familial and sporadic Alzheimer's disease
Saunders, A. M., Strittmatter, W. J., Schmechel, D., St. George-Hyslop, P. H., Pericak-Vance, M. A., Joo, S. H., Rosi, B. L., Gusella, J. F., Crapper-MacLachlan, D. R., Alberts, M. J., et al. (1993) · Neurology · doi:10.1212/wnl.43.8.1467 · Cited by 3,786
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Quantum-dot-tagged microbeads for multiplexed optical coding of biomolecules
Han, M., Gao, X., Su, J. Z., & Nie, S. (2001) · Nature Biotechnology · doi:10.1038/90228 · Cited by 2,631
The power and promise of population genomics: from genotyping to genome typing
Luikart, G., England, P. R., Tallmon, D., Jordan, S., & Taberlet, P. (2003) · Nature Reviews Genetics · doi:10.1038/nrg1226 · Cited by 1,365
Accessing genetic variation: genotyping single nucleotide polymorphisms
Syvänen, A.-C. (2001) · Nature Reviews Genetics · doi:10.1038/35103535 · Cited by 1,083
Dunbar, S. A. (2006) · Clinica Chimica Acta · doi:10.1016/j.cccn.2005.06.023 · Cited by 608
Methods for Genotyping Single Nucleotide Polymorphisms
Kwok, P.-Y. (2001) · Annual Review of Genomics and Human Genetics · doi:10.1146/annurev.genom.2.1.235 · Cited by 580
Metadata
- Authors
- Chen, J., Iannone, M. A., Li, M.-S., Taylor, J. D., Rivers, P., Nelsen, A. J., Slentz-Kesler, K. A., Roses, A., & Weiner, M. P.
- Venue
- Genome Research
- Publisher
- Cold Spring Harbor Laboratory
- Published
- 2000-04-01
- Volume
- 10
- Issue
- 4
- Pages
- 549-557
- ISSN
- 1088-9051 (ISSN-L)
- 1549-5469
- OpenAlex
- W1943638243
- Cited by
- 211
- References
- 1
- Type
- posted content
- Language
- en
- Open access
- bronze
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- Chen, J., Iannone, M. A., Li, M.-S., Taylor, J. D., Rivers, P., Nelsen, A. J., Slentz-Kesler, K. A., Roses, A., & Weiner, M. P. (2000). A Microsphere-Based Assay for Multiplexed Single Nucleotide Polymorphism Analysis Using Single Base Chain Extension. Genome Research. https://doi.org/10.1101/gr.10.4.549
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