OligoY pipeline for full Y chromosome painting

Abstract

AbstractMotivationThe standard protocol for designing probes used in full chromosome fluorescent labeling experiments does not include repetitive sequences to avoid off-target hybridization. Due to the Y chromosome’s highly repetitive nature, most assembly nowadays still have heavily fragmented and incomplete Y sequences. Among these, the remaining non-repetitive sequences are insufficient to design probes and efficiently perform FISH Oligopaint assays, since they do not cover most regions of the chromosome. Ergo, cytogenetic studies with the Y are sparse, and analysis such as its function throughout the cell cycle and insights into its evolutionary history and relationships with other regions of the genome remain poorly studied.ResultsIn this work, we introduce a new pipeline for designing FISH Oligopaint probes for the Y chromosome of any species of interest. OligoY pipeline uses open-source tools, enriches the amount of contigs assigned to the Y chromosome from the draft assembly, and effectively uses repetitive sequences unique to the target chromosome to design probes. Throughout all of its steps, the pipeline guarantees the user the autonomy to choose parameters, thus maximizing overall efficiency of cytogenetic experiments. After extensive in silico andin situtests and validations withDrosophila melanogaster, we showed for the first time a pipeline for probe design that significantly increases previous Y chromosome staining with no off-target signal.AvailabilityThe pipeline is available athttps://github.com/isabela42/OligoY.