Background: The development of long-read sequencing technologies, such as single-molecule real-time (SMRT) sequencing by PacBio, has produced a revolution in the sequencing of small genomes. Sequencing organelle genomes using PacBio long-read data is a cost effective, straightforward approach. Nevertheless, the availability of simple-to-use software to perform the assembly from raw reads is limited at present.
Results: We present Organelle-PBA, a Perl program designed specifically for the assembly of chloroplast and mitochondrial genomes. For chloroplast genomes, the program selects the chloroplast reads from a whole genome sequencing pool, maps the reads to a reference sequence from a closely related species, and then performs read correction and de novo assembly using Sprai. Organelle-PBA completes the assembly process with the additional step of scaffolding by SSPACE-LongRead. The program then detects the chloroplast inverted repeats and reassembles and re-orients the assembly based on the organelle origin of the reference. We have evaluated the performance of the software using PacBio reads from different species, read coverage, and reference genomes. Finally, we present the assembly of two novel chloroplast genomes from the species Picea glauca (Pinaceae) and Sinningia speciosa (Gesneriaceae).
Conclusion: Organelle-PBA is an easy-to-use Perl-based software pipeline that was written specifically to assemble mitochondrial and chloroplast genomes from whole genome PacBio reads. The program is available at https://github.com/aubombarely/Organelle_PBA.
Digital Object Identifier (DOI)
The program and the experiments reported in this article were funded with start-up funds provided to Dr. Bombarely at Virginia Tech (Program: Plant and Genomic Cluster).
The datasets supporting the conclusions in this article are available in the NCBI SRA repository:
For Arabidopsis thaliana:
For Mus musculus:
For Picea glauca:
Soorni, Aboozar; Haak, David; Zaitlin, David; and Bombarely, Aureliano, "Organelle_PBA, a Pipeline for Assembling Chloroplast and Mitochondrial Genomes from PacBio DNA Sequencing Data" (2017). Kentucky Tobacco Research and Development Center Faculty Publications. 19.