Background: Plastid genomes typically display a circular, quadripartite structure with two inverted repeat regions, which challenges automatic assembly procedures. The correct assembly of plastid genomes is a prerequisite for the validity of subsequent analyses on genome structure and evolution. The average coverage depth of a genome assembly is often used as an indicator of assembly quality. Visualizing coverage depth across a draft genome is a critical step, which allows users to inspect the quality of the assembly and, where applicable, identify regions of reduced assembly confidence. Despite the interplay between genome structure and assembly quality, no contemporary, user-friendly software tool can visualize the coverage depth of a plastid genome assembly while taking its quadripartite genome structure into account. A software tool is needed that fills this void. Results: We introduce 'PACVr', an R package that visualizes the coverage depth of a plastid genome assembly in relation to the circular, quadripartite structure of the genome as well as the individual plastome genes. By using a variable window approach, the tool allows visualizations on different calculation scales. It also confirms sequence equality of, as well as visualizes gene synteny between, the inverted repeat regions of the input genome. As a tool for plastid genomics, PACVr provides the functionality to identify regions of coverage depth above or below user-defined threshold values and helps to identify non-identical IR regions. To allow easy integration into bioinformatic workflows, PACVr can be invoked from a Unix shell, facilitating its use in automated quality control. We illustrate the application of PACVr on four empirical datasets and compare visualizations generated by PACVr with those of alternative software tools. Conclusions: PACVr provides a user-friendly tool to visualize (a) the coverage depth of a plastid genome assembly on a circular, quadripartite plastome map and in relation to individual plastome genes, and (b) gene synteny across the inverted repeat regions. It contributes to optimizing plastid genome assemblies and increasing the reliability of publicly available plastome sequences. The software, example datasets, technical

Background:The circular, quadripartite structure of plastid genomes which includes two inverted repeat regions renders the automatic assembly of plastid genomes challenging. The correct assembly of plastid genomes is a prerequisite for the validity of subsequent analyses on plastid genome structure and evolution. Plastome-based phylogenetic or population genetic investigations, for example, require the precise identification of DNA sequence and length to determine the location of nucleotide polymorphisms. The average coverage depth of a genome assembly is often used as an indicator for assembly quality. Visualizing coverage depth across a draft genome allows users to inspect the quality of the assembly and, where applicable, identify regions of reduced assembly confidence. Based on such visualizations, users can conduct a local re-assembly or other forms of targeted error correction. Few, if any, contemporary software tools can visualize the coverage depth of a plastid genome assembly while taking its quadripartite structure into account, despite the interplay between genome structure and assembly quality. A software tool is needed that visualizes the coverage depth of a plastid genome assembly on a circular, quadripartite map of the plastid genome. Results:We introduce 'PACVr', an R package that visualizes the coverage depth of a plastid genome assembly in relation to the circular, quadripartite structure of the genome as well as to the individual plastome genes. The tool allows visualizations on different scales using a variable window approach and also visualizes the equality of gene synteny in the inverted repeat regions of the plastid genome, thus providing an additional measure of assembly quality. As a tool for plastid genomics, PACVr provides the functionality to identify regions of coverage depth above or below user-defined threshold values and helps to identify non-identical IR regions. To allow easy integration into bioinformatic workflows, PACVr can be directly invoked from a Unix shell, thus facilitating its use in automated quality control. We illustrate the application of PACVr on two empirical datasets and compare the resulting visualizations with alternative software tools for displaying plastome sequencing coverage. Conclusions:PACVr provides a user-friendly tool to visualize (a) the coverage depth of a plastid genome assembly on a circular, quadripartite plastome map and in relation to individual plastome genes, and (b) the equality of gene synteny in the inverted repeat regions. It, thus, contributes to optimizing plastid genome assemblies and increasing the reliability of publicly available plastome sequences, especially in light of incongruence among the visualization results of alternative software tools. The software, example datasets, technical documentation, and a tutorial are available with the package at https://github.com/michaelgruenstaeudl/PACVr.

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In plastid genomes, the depth and evenness of sequence coverage are considered important indicators for assembly quality. However, the precise manifestations that sequencing depth and evenness can have in the assembly of these genomes, as well as any differences across individual genome sections, have yet to be evaluated. This investigation aims to identify the impact that sequencing depth and evenness can have on the assembly of plastid genomes and how both metrics are related to plastid genome structure. Specifically, we assess if sequencing evenness and reduced sequencing depth have significant correlations with, or significant differences among, individual genome sections, assembly quality metrics, the sequencing platforms employed, and the software tools used for genome assembly. To that end, we retrieve published plastid genomes as well as their sequence reads and genome metadata from public databases, measure sequencing depth and evenness across their sequences, and test several hypotheses on genome assembly and structure through non-parametric statistical tests. The results of our analyses show significant differences in sequencing depth across the four structural partitions as well as between the coding and non-coding sections of the plastid genomes, a significant correlation between sequencing evenness and the number of ambiguous nucleotides per genome, and significant differences in sequencing evenness between various sequencing platforms. Based on these results, we conclude that the observed differences and correlations are not a product of chance alone but possibly genuine manifestations of sequencing depth and evenness during the assembly of these genomes.

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