Assembly and alignment
Mitogenome amplicon reads were quality filtered with Trimmomatic v.0.33
(Bolger, Lohse, and Usadel 2014) with parameters SLIDINGWINDOW: 4:15 and
MINLEN: 36. Since the only publicly available mitogenome representing
any Tupaia species (the northern treeshrew, T. belangeriNC_002521; Schmitz, Ohme, & Zischler, 2000) is highly divergent from
our study species (Roberts et al., 2011), we first generated reference
mitogenomes for the mountain treeshrew and 3 more closely related
outgroup species: the pygmy treeshrew, large treeshrew, and ruddy
treeshrew. For each species, we selected one individual with the highest
number of sequencing reads (pygmy treeshrew, BOR 443; large treeeshrew,
BOR 010, & ruddy treeshrew, UMMZ174429) and assembled sequencesde novo using the MIRA v1.0.1 plugin in Geneious v9.1.2
(Biomatters Ltd.). Quality filtered sequence reads were mapped to the
appropriate reference using BWA-MEM v0.7.10 (Li 2013) with default
parameters. We also assembled mitogenomes from UCE-enriched library
sequences (Supplemental Information). Consensus sequences were generated
with Geneious (lowest coverage to call a base 5×, and Highest Total
Quality parameters) and aligned with the MAFFT v7.450 plugin (Katoh et
al. 2002). We transferred annotations from the northern treeshrew
reference to the consensus sequences. To rule out the presence of
nuclear copies of mitochondrial genes (NUMTs), we translated all
protein-coding genes to check for frame shifts or stop codons.