Abstract
A current strategy for obtaining haplotype information from several individuals involves short-read sequencing of pooled amplicons, where fragments from each individual is identified by a unique DNA barcode. In this paper, we report a new method to recover the phylogeny of haplotypes from short-read sequences obtained using pooled amplicons from a mixture of individuals, without barcoding. The method, AFPhyloMix, accepts an alignment of the mixture of reads against a reference sequence, obtains the single-nucleotide-polymorphisms (SNP) patterns along the alignment, and constructs the phylogenetic tree according to the SNP patterns. AFPhyloMix adopts a Bayesian inference model to estimate the phylogeny of the haplotypes and their relative abundances, given that the number of haplotypes is known. In our simulations, AFPhyloMix achieved at least 80% accuracy at recovering the phylogenies and relative abundances of the constituent haplotypes, for mixtures with up to 15 haplotypes. AFPhyloMix also worked well on a real data set of kangaroo mitochondrial DNA sequences.
Highlights
Molecular phylogenetic reconstruction is the mainstay of modern evolutionary biology [1, 2]
This research demonstrates the feasibility of reconstructing a phylogenetic tree directly from the short read sequences obtained from a mixture of closely related amplified sequences, without barcoding
AFPhyloMix is designed to estimate the concentration of haplotypes and reconstruct the phylogenetic tree directly from the short read sequences in the mixture of haplotypes with no barcode, given that the number of haplotypes is known
Summary
Molecular phylogenetic reconstruction is the mainstay of modern evolutionary biology [1, 2]. Because modern sequencing technologies can produce several gigabases of nucleotide sequences in a single day, one of the challenges for the molecular phylogeneticist is to deal with this quantity of data in a timely manner while still reconstructing accurate phylogenies To this end, phylogeneticists have developed rapid alignment and tree reconstruction algorithms [4, 5], using pre-processed and curated sequences. Pre-processing and sequence curation can be laborious, but are necessary tasks because a great deal of sequence data are generated using generation short-read sequencing technologies Such sequences are often barcoded using unique DNA identifier tags, and collectively pooled and sequenced in a single run. The unique barcode allows sequences belonging to different samples to be separated computationally, before additional error-correction and subsequent down-stream analyses are performed
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