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. 2014 May;42(9):e80.
doi: 10.1093/nar/gku210. Epub 2014 Mar 14.

RepARK--de novo creation of repeat libraries from whole-genome NGS reads

Philipp Koch  1 , Matthias Platzer  2 , Bryan R Downie  2
Affiliations

RepARK--de novo creation of repeat libraries from whole-genome NGS reads

Philipp Koch et al. Nucleic Acids Res. 2014 May.

Abstract

Generation of repeat libraries is a critical step for analysis of complex genomes. In the era of next-generation sequencing (NGS), such libraries are usually produced using a whole-genome shotgun (WGS) derived reference sequence whose completeness greatly influences the quality of derived repeat libraries. We describe here a de novo repeat assembly method--RepARK (Repetitive motif detection by Assembly of Repetitive K-mers)--which avoids potential biases by using abundant k-mers of NGS WGS reads without requiring a reference genome. For validation, repeat consensuses derived from simulated and real Drosophila melanogaster NGS WGS reads were compared to repeat libraries generated by four established methods. RepARK is orders of magnitude faster than the other methods and generates libraries that are: (i) composed almost entirely of repetitive motifs, (ii) more comprehensive and (iii) almost completely annotated by TEclass. Additionally, we show that the RepARK method is applicable to complex genomes like human and can even serve as a diagnostic tool to identify repetitive sequences contaminating NGS datasets.

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Figures

Figure 1.
Figure 1.
Workflow of the repeat library creation pipeline RepARK. WGS sequencing reads (a) contain unique (black) and repetitive (red) fractions of the genome. K-mers of all reads (b) were counted and the threshold of frequent k-mers is determined. These abundant k-mers are isolated (c) and assembled by a de novo genome assembly program (such as Velvet) into repeat consensus sequences (d).
Figure 2.
Figure 2.
Cumulative length of repetitive and non-repetitive consensuses within each library. Black: repetitive consensuses (i.e. align more than once to the reference); gray: non-repetitive consensuses (i.e. singly mapping or not at all); Sanger: libraries based on Sanger sequencing data; simulated: libraries derived from simulated NGS reads; real: libraries derived from Illumina reads.
Figure 3.
Figure 3.
Repeat fractions identified in the D. melanogaster reference sequence. Black: fraction of the reference masked by RepeatMasker using the respective repeat library; gray: fraction of the reference that was subsequently masked by RepeatMasker using RepBase; Sanger: libraries based on Sanger sequencing data; simulated: libraries derived from simulated NGS reads; real: libraries derived from Illumina reads.
Figure 4.
Figure 4.
Boxplot of DmRepBase repeat class completeness in the de novo repeat libraries. DNA: 33 DNA transposons; LTR: 138 LTR retrotransposons; non-LTR: 41 non-LTR retrotransposons; Sanger: libraries based on Sanger sequencing data; simulated: libraries derived from simulated NGS reads; real: libraries derived from Illumina reads; box: first and third quartiles; horizontal line: median; whiskers: most extreme value within 1.5× of inter-quartile range; dots: outliers. A full table of repeat family representation in the RepARK libraries can be found in Supplementary Table S3.
Figure 5.
Figure 5.
Fractions of known D. melanogaster segmental duplications identified by the de novo repeat libraries. Sanger: libraries based on Sanger sequencing data; simulated: libraries derived from simulated NGS reads; real: libraries derived from Illumina reads.
Figure 6.
Figure 6.
Fractions of the D. melanogaster genome reference classified according to annotated repeat libraries. Black: DNA transposon sequence; dark gray: retrotransposon sequence; light gray: unclear; Sanger: libraries based on Sanger sequencing data; simulated: libraries derived from simulated NGS reads; real: libraries derived from Illumina reads.
Figure 7.
Figure 7.
High confidence alignments of human RepARK consensuses (right half) to the Epstein-Barr virus genome (left half, HHV-4). Each ribbon represents a consensus alignment with >90% mapping and p < 10−60, encompassing 90.5% of the Epstein-Barr virus genome. Lower confidence consensuses align to the remaining 9.5% with more relaxed criteria. Three consensuses map multiple times to the virus genome sequence (NODE_48265, NODE_888, NODE_5085; dark red). Created with Circoletto (http://bat.ina.certh.gr/tools/circoletto/).
See this image and copyright information in PMC

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