title: In-solution hybridization for mammalian mitogenome enrichment: pros, cons and challenges associated with multiplexing degraded DNA.
abstract: Here, we present a set of RNA-based probes for whole mitochondrial genome in-solution enrichment, targeting a diversity of mammalian mitogenomes. This probes set was designed from seven mammalian orders and tested to determine the utility for enriching degraded DNA. We generated 63 mitogenomes representing five orders and 22 genera of mammals that yielded varying coverage ranging from 0 to >5400X. Based on a threshold of 70% mitogenome recovery and at least 10× average coverage, 32 individuals or 51% of samples were considered successful. The estimated sequence divergence of samples from the probe sequences used to construct the array ranged up to nearly 20%. Sample type was more predictive of mitogenome recovery than sample age. The proportion of reads from each individual in multiplexed enrichments was highly skewed, with each pool having one sample that yielded a majority of the reads. Recovery across each mitochondrial gene varied with most samples exhibiting regions with gaps or ambiguous sites. We estimated the ability of the probes to capture mitogenomes from a diversity of mammalian taxa not included here by performing a clustering analysis of published sequences for 100 taxa representing most mammalian orders. Our study demonstrates that a general array can be cost and time effective when there is a need to screen a modest number of individuals from a variety of taxa. We also address the practical concerns for using such a tool, with regard to pooling samples, generating high quality mitogenomes and detail a pipeline to remove chimeric molecules.
authors.affiliation: Center for Conservation and Evolutionary Genetics, Smithsonian Conservation Biology Institute, National Zoological Park, Washington, DC, 20008, USA.|Division of Mammals, National Museum of Natural History, MRC 108, Smithsonian Institution, PO Box 37012, Washington, DC, 20013-7012, USA.|Department of Environmental Science & Policy, George Mason University, Fairfax, VA, 22030, USA.
authors.forename: Melissa T R
authors.initials: MT
authors.lastname: Hawkins
authors.affiliation: Center for Conservation and Evolutionary Genetics, Smithsonian Conservation Biology Institute, National Zoological Park, Washington, DC, 20008, USA.|Program in Human Ecology and Archaeobiology, Department of Anthropology, National Museum of Natural History, Smithsonian Institution, PO Box 37012, Washington, DC, 20013-7012, USA.|Department of Anthropology, University of Maryland, College Park, MD, 20742, USA.
authors.forename: Courtney A
authors.initials: CA
authors.lastname: Hofman
authors.affiliation: Center for Conservation and Evolutionary Genetics, Smithsonian Conservation Biology Institute, National Zoological Park, Washington, DC, 20008, USA.|Department of Animal & Avian Sciences, University of Maryland, College Park, MD, 20742, USA.
authors.forename: Taylor
authors.initials: T
authors.lastname: Callicrate
authors.affiliation: Center for Conservation and Evolutionary Genetics, Smithsonian Conservation Biology Institute, National Zoological Park, Washington, DC, 20008, USA.|Division of Mammals, National Museum of Natural History, MRC 108, Smithsonian Institution, PO Box 37012, Washington, DC, 20013-7012, USA.
authors.forename: Molly M
authors.initials: MM
authors.lastname: McDonough
authors.affiliation: Center for Conservation and Evolutionary Genetics, Smithsonian Conservation Biology Institute, National Zoological Park, Washington, DC, 20008, USA.|Division of Mammals, National Museum of Natural History, MRC 108, Smithsonian Institution, PO Box 37012, Washington, DC, 20013-7012, USA.|Department of Environmental Science & Policy, George Mason University, Fairfax, VA, 22030, USA.
authors.forename: Mirian T N
authors.initials: MT
authors.lastname: Tsuchiya
authors.affiliation: Center for Conservation and Evolutionary Genetics, Smithsonian Conservation Biology Institute, National Zoological Park, Washington, DC, 20008, USA.|Division of Mammals, National Museum of Natural History, MRC 108, Smithsonian Institution, PO Box 37012, Washington, DC, 20013-7012, USA.
authors.forename: Eliécer E
authors.initials: EE
authors.lastname: Gutiérrez
authors.affiliation: Division of Mammals, National Museum of Natural History, MRC 108, Smithsonian Institution, PO Box 37012, Washington, DC, 20013-7012, USA.
authors.forename: Kristofer M
authors.initials: KM
authors.lastname: Helgen
authors.affiliation: Center for Conservation and Evolutionary Genetics, Smithsonian Conservation Biology Institute, National Zoological Park, Washington, DC, 20008, USA.|Division of Mammals, National Museum of Natural History, MRC 108, Smithsonian Institution, PO Box 37012, Washington, DC, 20013-7012, USA.
authors.forename: Jesus E
authors.initials: JE
authors.lastname: Maldonado
mesh_terms: D000818:Animals; D016000:Cluster Analysis; D004272:DNA, Mitochondrial / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification*; D008322:Mammals / Q000235:genetics*; D009693:Nucleic Acid Hybridization / Q000379:methods*; D015345:Oligonucleotide Probes / Q000235:genetics*; D017422:Sequence Analysis, DNA
chemical_list: D004272:DNA, Mitochondrial; D015345:Oligonucleotide Probes
keywords: high throughput sequencing; mitogenome; museomics; targeted sequence capture