Computational protocol: Chromoanasynthetic Genomic Rearrangement Identified in a N-Ethyl-N-Nitrosourea (ENU) Mutagenesis Screen in Caenorhabditis elegans

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Protocol publication

[…] Paired-end sequence reads were mapped to the C. elegans reference genome version WS230 (www.wormbase.org) using both short-read aligners BWA () and Phaster (Philip Green, personal communication). The resulting alignment files were sorted and indexed, and single nucleotide variants (SNVs) were identified with the help of the SAMtools toolbox (). Copy numbers were estimated in a given genomic interval by dividing the number of aligned reads for strain BQ13 by the number of reads in the corresponding interval for the parental strain, after proper rescaling to a common total number of aligned reads for each library. The estimated copy number in overlapping intervals was examined visually using R (www.r-project.org), and the size of the intervals was varied in order to find the approximate location of each breakpoint. The creation of those overlapping intervals and the calculation of the number of reads within each interval were performed with the BEDTools suite (). Using the IGV genome viewer (; ), read alignments around the approximate breakpoints were examined for multiple split-reads with alignments ending at the same location, with the second part of those reads all starting to align at a common location and orientation. In order to confirm the exact breakpoints and junctions found in IGV, a subset of split reads overlapping the junctions were realigned on the reference genome using Blast () as implemented on WormBase (www.wormbase.org). […]

Pipeline specifications

Software tools BWA, PHASTER, SAMtools, BEDTools, IGV
Databases WormBase
Applications Genome annotation, Genome data visualization
Organisms Caenorhabditis elegans, Homo sapiens
Chemicals Ethylnitrosourea