title: Assessing the utility of whole genome amplified DNA for next-generation molecular ecology.
abstract: DNA quantity can be a hindrance in ecological and evolutionary research programmes due to a range of factors including endangered status of target organisms, available tissue type, and the impact of field conditions on preservation methods. A potential solution to low-quantity DNA lies in whole genome amplification (WGA) techniques that can substantially increase DNA yield. To date, few studies have rigorously examined sequence bias that might result from WGA and next-generation sequencing of nonmodel taxa. To address this knowledge deficit, we use multiple displacement amplification (MDA) and double-digest RAD sequencing on the grey mouse lemur (Microcebus murinus) to quantify bias in genome coverage and SNP calls when compared to raw genomic DNA (gDNA). We focus our efforts in providing baseline estimates of potential bias by following manufacturer's recommendations for starting DNA quantities (>100 ng). Our results are strongly suggestive that MDA enrichment does not introduce systematic bias to genome characterization. SNP calling between samples when genotyping both de-novo and with a reference genome are highly congruent (>98%) when specifying a minimum threshold of 20X stack depth to call genotypes. Relative genome coverage is also similar between MDA and gDNA, and allelic dropout is not observed. SNP concordance varies based on coverage threshold, with 95% concordance reached at ~12X coverage genotyping de-novo and ~7X coverage genotyping with the reference genome. These results suggest that MDA may be a suitable solution for next-generation molecular ecological studies when DNA quantity would otherwise be a limiting factor.
authors.affiliation: Department of Biology, Duke University, Box 90338, BioSci 130 Science Drive, Durham, NC, 27708, USA.
authors.forename: Christopher
authors.initials: C
authors.lastname: Blair
authors.affiliation: Department of Biology, Duke University, Box 90338, BioSci 130 Science Drive, Durham, NC, 27708, USA.
authors.forename: C Ryan
authors.initials: CR
authors.lastname: Campbell
authors.affiliation: Department of Biology, Duke University, Box 90338, BioSci 130 Science Drive, Durham, NC, 27708, USA.
authors.forename: Anne D
authors.initials: AD
authors.lastname: Yoder
mesh_terms: D000818:Animals; D016565:Cheirogaleidae / Q000235:genetics; D023281:Genomics / Q000379:methods*; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D020641:Polymorphism, Single Nucleotide
keywords: biotechnology; environmental DNA; genetics; genomics; landscape; phylogeography; proteomics