Abstract
Emulsion-fusion PCR recovers long-range sequence information by combining products in cis from individual genomic DNA molecules. Emulsion droplets act as very numerous small reaction chambers in which different PCR products from a single genomic DNA molecule are condensed into short joint products, to unite sequences in cis from widely separated genomic sites. These products can therefore provide information about the arrangement of sequences and variants at a larger scale than established long-read sequencing methods. The method has been useful in defining the phase of variants in haplotypes, the typing of inversions, and determining the configuration of sequence variants in multiallelic CNVs. In this description we outline the rationale for the application of emulsion-fusion PCR methods to the analysis of multiallelic CNVs, and give practical details for our own implementation of the method in that context.
This is a preview of subscription content, log in via an institution.
Buying options
Tax calculation will be finalised at checkout
Purchases are for personal use only
Learn about institutional subscriptionsReferences
Hastie AR, Dong L, Smith A et al (2013) Rapid genome mapping in nanochannel arrays for highly complete and accurate de novo sequence assembly of the complex Aegilops tauschii genome. PLoS One 8:e55864
Duitama J, McEwen GK, Huebsch T et al (2012) Fosmid-based whole genome haplotyping of a HapMap trio child: evaluation of Single Individual Haplotyping techniques. Nucleic Acids Res 40:2041–2053
Suk E-K, McEwen GK, Duitama J et al (2011) A comprehensively molecular haplotype-resolved genome of a European individual. Genome Res 21:1672–1685
Huddleston J, Ranade S, Malig M et al (2014) Reconstructing complex regions of genomes using long-read sequencing technology. Genome Res 24:688–696
Cherf GM, Lieberman KR, Rashid H et al (2012) Automated forward and reverse ratcheting of DNA in a nanopore at 5-A precision. Nat Biotechnol 30:344–348
Wetmur JG, Chen J (2011) Linking emulsion PCR haplotype analysis. Methods Mol Biol 687:165–175
Wetmur JG, Kumar M, Zhang L et al (2005) Molecular haplotyping by linking emulsion PCR: analysis of paraoxonase 1 haplotypes and phenotypes. Nucleic Acids Res 33:2615–2619
Turner DJ, Shendure J, Porreca G et al (2006) Assaying chromosomal inversions by single-molecule haplotyping. Nat Methods 3:439–445
Turner DJ, Tyler-Smith C, Hurles ME (2008) Long-range, high-throughput haplotype determination via haplotype-fusion PCR and ligation haplotyping. Nucleic Acids Res 36:e82
Turner DJ, Hurles ME (2009) High-throughput haplotype determination over long distances by haplotype fusion PCR and ligation haplotyping. Nat Protoc 4:1771–1783
Black H, Khan F, Tyson J et al (2014) Inferring mechanisms of copy number change from haplotype structures at the human DEFA1A3 locus. BMC Genomics 15:614
Tyson J, Armour JAL (2012) Determination of haplotypes at structurally complex regions using emulsion haplotype fusion PCR. BMC Genomics 13:693
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2017 Springer Science+Business Media New York
About this protocol
Cite this protocol
Tyson, J., Armour, J.A.L. (2017). Analysis of Multiallelic CNVs by Emulsion Haplotype Fusion PCR. In: White, S., Cantsilieris, S. (eds) Genotyping. Methods in Molecular Biology, vol 1492. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6442-0_10
Download citation
DOI: https://doi.org/10.1007/978-1-4939-6442-0_10
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-6440-6
Online ISBN: 978-1-4939-6442-0
eBook Packages: Springer Protocols