Genome editing tools have recently been developed that are capable of generating deletions and/or rearrangements in plant mitochondrial genomes. We utilized custom-designed meganuclease constructs targeting the essential mitochondrial gene atp1 in tobacco plants carrying a version of atp1 that had been repurposed to function as a nuclear gene. Leaf extracts enriched in mitochondria from three independent genome editing events were evaluated by PacBio sequencing and analysis. Contigs that display sequence homology to the reference mitochondrial genome BA000042 are deposited here. Some of the contigs correspond to the tobacco mitochondrial genome as expected, while others corresponded to sequences from the nucleus where the ancient transfer of portions of the mitochondrial genome became incorporated into the nuclear genome. Still other contigs correspond to the chloroplast genome, or pieces of the chloroplast genome anciently transferred to the nucleus, as small fragments of the chloroplast genome are interspersed in plant mitochondrial genomes. Analysis of contigs of genuine mitochondrial origin demonstrated that the double-strand breaks imposed by the genome editing enzymes resulted in the deletion of atp1 and neighboring sequences, along with rearrangements in overall genome structure.

Three tobacco samples that have been enriched for the mitochondria (MT) were sequenced on a PacBio Sequel IIe in HiFi-CCS mode. Sequences in the vicinity of the essential gene atp1 in the mitochondrial genomes have been edited.

The HiFi reads for the three samples were mapped to reference tobacco MT genome BA000042 using Minimap2 (version 2.24-r1122) with the flags -ax map-hifi --sam-hit-only. The read identifiers for the reads that mapped to BA000042 were then saved and used to select a subset of the raw reads. The read subset for each sample was assembled using hifiasm (version 0.16.1-r375) with the -l 0 flag and the assembled contigs can be found in the fasta files. In select cases, trimmed, hand-curated contigs are also shown in addition to the original assembly fasta file.

The data files are in fasta format.