Plastid Genome Assembly Using Long-read data (ptGAUL)

Abstract

Although plastid genome (plastome) structure is highly conserved across most seed plants, investigations during the past two decades have revealed several disparately related lineages that have experienced substantial rearrangements. Most plastomes have two inverted repeat regions and two single-copy regions with few dispersed repeats. However, the plastomes of some taxa do harbor long repeat sequences (>300 bp). These long repeats make it difficult to assemble complete plastomes using short read data, leading to misassemblies and consensus sequences that have spurious rearrangements. Long read sequencing can potentially overcome these challenges. However, there is no consensus as to the most effective method for accurately assembling plastomes using long read data. Here, we generated a pipeline, plastid Genome Assembly Using Long-read data (ptGAUL) to address the problem of assembling of plastomes using long read data from Oxford Nanopore Technologies (ONT) or Pacific Biosciences (Pacbio) platforms. We demonstrated the efficacy of the ptGAUL pipeline using 16 published long read datasets. We showed that ptGAUL produces accurate and unbiased assemblies. Additionally, we applied ptGAUL to assemble four Juncus (Juncaceae) plastomes using ONT long reads. Our results revealed many long repeats and rearrangements in Juncus plastomes compared with basal lineages of Poales.

Chloroplast genome‐based genetic resources via genome skimming for the subalpine forests of Japan and adjacent regions

“…Instead reads were mapped to reference chloroplast genomes with 'bwa mem' (Li, 2013). Vaccinium vitis-idaea reads were mapped to a V. vitis-idaea whole chloroplast genome (sourced from a plant cultivated in Canada, DNA Data Bank of Japan accession number: BR002395) assembled from Oxford Nanopore (SRR25468450) and Illumina NovaSeq reads (SRR25477290) (Hirabayashi et al, 2023) using the ptGAUL pipeline (Zhou et al, 2023) (https:// github. com/ Bean0 61/ ptgaul).…”

The complete chloroplast genome of Marupa (Simarouba amara Aubl., Simaroubaceae)

“…Chloroplast genome assembly was done using the pipeline pt-GAUL 1.0.5 (Zhou et al, 2023), using default parameters. Briefly, long-reads were mapped using Minimap2 (Li, 2018) to the reference chloroplast genome of Ailanthus altissima (MG799542.1)…”

Chloroplast genome-based genetic resources for Japan’s threatened subalpine forests via genome skimming

“…Instead reads were mapped to reference chloroplast genomes with 'bwa mem' (Li, 2013). Vaccinium vitis-idaea reads were mapped to a V. vitis-idaea whole chloroplast genome (sourced from a plant cultivated in Canada) assembled from Oxford Nanopore (SRR25468450) and Illumina NovaSeq reads (SRR25477290) (Hirabayashi, Debnath and Owens, 2023) using the ptGAUL pipeline (Zhou et al, 2023) (https://github.com/Bean061/ptgaul). Before chloroplast genome assembly, Nanopore reads were filtered by Filtlong (https://github.com/rrwick/Filtlong) with parameters of '--min_length 1000 --keep_percent 90,' while Illumina reads (1 Gbp for each pair) were down-sampled by the 'sample' subcommand of seqtk (https://github.com/lh3/seqtk) ver.…”