title: Impacts of low coverage depths and post-mortem DNA damage on variant calling: a simulation study.
abstract: BACKGROUND
  Massively parallel sequencing platforms, featuring high throughput and relatively short read lengths, are well suited to ancient DNA (aDNA) studies. Variant identification from short-read alignment could be hindered, however, by low DNA concentrations common to historic samples, which constrain sequencing depths, and post-mortem DNA damage patterns.
  
  
  RESULTS
  We simulated pairs of sequences to act as reference and sample genomes at varied GC contents and divergence levels. Short-read sequence pools were generated from sample sequences, and subjected to varying levels of "post-mortem" damage by adjusting levels of fragmentation and fragmentation biases, transition rates at sequence ends, and sequencing depths. Mapping of sample read pools to reference sequences revealed several trends, including decreased alignment success with increased read length and decreased variant recovery with increased divergence. Variants were generally called with high accuracy, however identification of SNPs (single-nucleotide polymorphisms) was less accurate for high damage/low divergence samples. Modest increases in sequencing depth resulted in rapid gains in total variant recovery, and limited improvements to recovery of heterozygous variants.
  
  
  CONCLUSIONS
  This in silico study suggests aDNA-associated damage patterns minimally impact variant call accuracy and recovery from short-read alignment, while modest increases in sequencing depth can greatly improve variant recovery.
authors.affiliation: Environmental Futures Research Institute, Griffith University, Nathan, 4111, Queensland, Australia. m.parks@griffith.edu.au.
authors.forename: Matthew
authors.initials: M
authors.lastname: Parks
authors.affiliation: Environmental Futures Research Institute, Griffith University, Nathan, 4111, Queensland, Australia. d.lambert@griffith.edu.au.
authors.forename: David
authors.initials: D
authors.lastname: Lambert
mesh_terms: D001482:Base Composition; D019295:Computational Biology; D003198:Computer Simulation; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D004249:DNA Damage / Q000235:genetics*; D059014:High-Throughput Nucleotide Sequencing; D020641:Polymorphism, Single Nucleotide; D017422:Sequence Analysis, DNA
chemical_list: D004247:DNA