A rapid molecular approach for chromosomal phasing.
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Abstract | Determining the chromosomal phase of pairs of sequence variants - the arrangement of specific alleles as haplotypes - is a routine challenge in molecular genetics. Here we describe Drop-Phase, a molecular method for quickly ascertaining the phase of pairs of DNA sequence variants (separated by 1-200 kb) without cloning or manual single-molecule dilution. In each Drop-Phase reaction, genomic DNA segments are isolated in tens of thousands of nanoliter-sized droplets together with allele-specific fluorescence probes, in a single reaction well. Physically linked alleles partition into the same droplets, revealing their chromosomal phase in the co-distribution of fluorophores across droplets. We demonstrated the accuracy of this method by phasing members of trios (revealing 100% concordance with inheritance information), and demonstrate a common clinical application by phasing CFTR alleles at genomic distances of 11-116 kb in the genomes of cystic fibrosis patients. Drop-Phase is rapid (requiring less than 4 hours), scalable (to hundreds of samples), and effective at long genomic distances (200 kb). |
Year of Publication | 2015
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Journal | PLoS One
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Volume | 10
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Issue | 3
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Pages | e0118270
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Date Published | 2015
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ISSN | 1932-6203
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DOI | 10.1371/journal.pone.0118270
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PubMed ID | 25739099
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PubMed Central ID | PMC4349636
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