Improved allelic differentiation using sequence-specific oligonucleotide hybridization incorporating an additional base-analogue mismatch.
Sequence-specific oligonucleotide hybridization (SSOH, 'dot-blotting') is a widely employed method of typing single nucleotide polymorphisms (SNPs), but it is often compromised by lack of allelic differentiation. We describe a novel improvement to SSOH that incorporates an additional misma...
| Main Authors: | , , , |
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| Format: | Journal article |
| Language: | English |
| Published: |
2004
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| Summary: | Sequence-specific oligonucleotide hybridization (SSOH, 'dot-blotting') is a widely employed method of typing single nucleotide polymorphisms (SNPs), but it is often compromised by lack of allelic differentiation. We describe a novel improvement to SSOH that incorporates an additional mismatch into the oligonucleotide probe using the universal base analogue 3-nitropyrrole. This method greatly increases allelic differentiation compared to standard SSOH where oligonucleotides contain only SNP-defining base changes. Moreover, stringency of the hybridisation is predictably maintained over a wide range of temperatures, which can be calculated empirically, thus facilitating the genotyping of multiple SNPs using similar conditions. This improved method increases the usefulness of hybridisation-based methods of rapid genotyping of SNPs and may have implications for array methodologies. |
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