Analyzing Copy Number Variation with Droplet Digital PCR

Avery Davis Bell; Christina L Usher; Steven A McCarroll

doi:10.1007/978-1-4939-7778-9_9

Analyzing Copy Number Variation with Droplet Digital PCR

Methods Mol Biol. 2018:1768:143-160. doi: 10.1007/978-1-4939-7778-9_9.

Authors

Avery Davis Bell^{1

2

3}, Christina L Usher^{1

2

3}, Steven A McCarroll^{4

5

6}

Affiliations

¹ Department of Genetics, Harvard Medical School, Boston, MA, USA.
² Program in Medical and Population Genetics, Broad Institute of MIT and Harvard, Cambridge, MA, USA.
³ Stanley Center for Psychiatric Research, Broad Institute of MIT and Harvard, Cambridge, MA, USA.
⁴ Department of Genetics, Harvard Medical School, Boston, MA, USA. mccarroll@genetics.med.harvard.edu.
⁵ Program in Medical and Population Genetics, Broad Institute of MIT and Harvard, Cambridge, MA, USA. mccarroll@genetics.med.harvard.edu.
⁶ Stanley Center for Psychiatric Research, Broad Institute of MIT and Harvard, Cambridge, MA, USA. mccarroll@genetics.med.harvard.edu.

PMID: 29717442
DOI: 10.1007/978-1-4939-7778-9_9

Abstract

Many genomic segments vary in copy number among individuals of the same species, or between cancer and normal cells within the same person. Correctly measuring this copy number variation is critical for studying its genetic properties, its distribution in populations and its relationship to phenotypes. Droplet digital PCR (ddPCR) enables accurate measurement of copy number by partitioning a PCR reaction into thousands of nanoliter-scale droplets, so that a genomic sequence of interest-whose presence or absence in a droplet is determined by end-point fluorescence-can be digitally counted. Here, we describe how we analyze copy number variants using ddPCR and review the design of effective assays, the performance of ddPCR with those assays, the optimization of reactions, and the interpretation of data.

Keywords: Copy number variants; Digital PCR; Droplet digital PCR; Genomic structural variation; Genotyping; Genotyping assay design.

Publication types

Research Support, N.I.H., Extramural

MeSH terms

Biological Variation, Individual
Biological Variation, Population / genetics
DNA / isolation & purification*
DNA Copy Number Variations / genetics*
Gene Dosage / genetics
Humans
Polymerase Chain Reaction / instrumentation
Polymerase Chain Reaction / methods*

Substances

DNA

Grants and funding

R01 HG006855/HG/NHGRI NIH HHS/United States