Full-Length Mitochondrial-DNA Sequencing on the PacBio RSII

Rolf H A M Vossen; Henk P J Buermans

doi:10.1007/978-1-4939-6442-0_12

Full-Length Mitochondrial-DNA Sequencing on the PacBio RSII

Methods Mol Biol. 2017:1492:179-184. doi: 10.1007/978-1-4939-6442-0_12.

Authors

Rolf H A M Vossen¹, Henk P J Buermans²

Affiliations

¹ Department of Human Genetics; Leiden Genome Technology Center, Leiden University Medical Center, Einthovenweg 20, 2333 ZC, Leiden, The Netherlands. r.vossen@lumc.nl.
² Department of Human Genetics; Leiden Genome Technology Center, Leiden University Medical Center, Einthovenweg 20, 2333 ZC, Leiden, The Netherlands.

PMID: 27822864
DOI: 10.1007/978-1-4939-6442-0_12

Abstract

Conventional mitochondrial-DNA (MT DNA) sequencing approaches use Sanger sequencing of 20-40 partially overlapping PCR fragments per individual, which is a time- and resource-consuming process. We have developed a high-throughput, accurate, fast, and cost-effective human MT DNA sequencing approach. In this setup we first generate long-range PCR products for two partially overlapping 7.7 and 9.2 kb MT DNA-specific amplicons, add sample-specific barcodes, and sequence these on the PacBio RSII system to obtain full-length MT DNA sequences for genotyping/haplotyping purposes.

Keywords: Haplotyping; Long read sequencing; Long-range PCR; MT-DNA; PacBio RSII; Sample multiplexing.

MeSH terms

DNA, Mitochondrial / genetics*
Humans
Polymerase Chain Reaction / methods*
Sequence Analysis, DNA / methods*

Substances

DNA, Mitochondrial