Unveiling the intricate structural variability induced by repeat-mediated recombination in the complete mitochondrial genome of Cuscuta gronovii Willd.

Abstract

Cuscuta gronovii Willd., a member of the Convolvulaceae family, is noted for its potential medicinal and nutritional benefits. In this study, we utilized a combination of Illumina and Oxford Nanopore sequencing technologies to successfully assemble the complete circular mitochondrial genome (mitogenome) of C. gronovii. The mitogenome, spanning 304,467 base pairs, includes 54 genes: 33 protein-coding genes, three ribosomal RNA (rRNA) genes, and 18 transfer RNA (tRNA) genes. Beyond its primary circular structure, we discovered and validated several alternative genomic conformations, driven by five specific repeat sequences. Three inverted repeats were found to initiate rearrangements, resulting in the creation of seven distinct chromosomal structures, while two direct repeats split a larger molecule into two subgenomic entities. We also mapped 421 RNA editing sites across the protein-coding sequences, influencing 33 protein-coding genes with varying distribution, particularly noting high frequencies in the nad4 and ccmB genes. Sixteen of these RNA editing sites were experimentally validated through PCR amplification and Sanger sequencing, confirming their presence with 100 % accuracy. This research not only introduces the first mitochondrial genome of C. gronovii but also highlights its complex conformational variability induced by repeat-mediated recombination, providing a valuable genomic resource for further molecular breeding efforts and phylogenetic evolution within the genus Cuscuta.