Case Study: Targeted high-Throughput Sequencing of Mitochondrial Genomes from Extinct Cave Bears via Direct Multiplex PCR Sequencing (DMPS)

Abstract

Here I describe the use of a recently developed technique for targeted high-throughput sequencing of highly degraded DNA by direct multiplex PCR sequencing (DMPS) that was used to amplify 31 near-complete mitochondrial genomes of the extinct cave bear (Ursus spelaeus). DMPS couples multiplex PCR with the generation of barcoded sequencing libraries to be sequenced in parallel on a high-throughput sequencing platform. DMPS makes it possible to generate large amounts of targeted DNA sequence data simultaneously from multiple degraded samples such as fossil remains. In this chapter, I describe an experiment that uses DMPS with different primer sets and on both modern and ancient DNA templates.

Note:

In the case study presented in this chapter, I describe the amplification and sequencing of whole mitochondrial genomes using a combined approach of the methods presented in Chapters 15 (12), and 19 (11). I discuss specific challenges associated with using this method to amplify and sequence modern and ancient DNA templates. For more information, see the original publication of the scientific results in Stiller et al. (2009) (4).