Assembly and comparative analysis of the complete mitochondrial genome of Tetrastigma Hemsleyanum Diels & Gilg: an invaluable traditional Chinese medicine

Tetrastigma hemsleyanum Diels & Gilg, a plant well-known in China for its use in traditional herbal medicine, is renowned for its remarkable medicinal value in various clinical contexts. Its chloroplast and nuclear genomes have been fully sequenced, providing valuable information for breeding and exploration. However, its mitochondrial genome remains uncharacterized, restricting the broader view of its evolution and its potential utilization. We present the assembled and annotated mitogenome of T. hemsleyanum to enhance the genetic resources for the Tetrastigma genus and pave the way for future in-depth studies of this medicinally significant plant. In this study, we assembled the mitochondrial genome of T. hemsleyanum var. “Jinhulu 2” by employing both the Illumina and Nanopore sequencing technologies and performed a comparative analysis with Vitis. The results revealed that the mitogenome is multi-chromosomal in structure, comprising two circular chromosomes with a total length of 809,173 bp and a GC content of 44.35%. It contains 76 annotated functional genes, including 35 protein-coding genes, 37 tRNA genes, 3 rRNA genes, and 1 pseudogene. Further analysis revealed 273 simple sequence repeats (SSRs), 109 tandem repeats, and 8,733 dispersed repeats, suggesting a higher repeat abundance in T. hemsleyanum compared to its closely related species. RNA editing analysis identified 57 sites within the mitochondrial CDS (coding DNA sequence), all showing a precise cytosine-to-uracil conversion, typically resulting in the substitution of hydrophilic amino acids with hydrophobic ones. The selection pressure analysis indicated that most mitochondrial CDS are under purifying selection, with only four genes (ccmB, ccmFc, ccmFn, and rps10) exhibiting positive selection. Notably, positive selection on ccmB was consistently observed across all pairwise species comparisons, highlighting its pivotal role in evolutionary adaptation. In addition, we identified 62 homologous fragments transferred from the chloroplast to the mitochondrial genome, including 15 intact chloroplast-derived tRNA genes. Comparative analysis revealed that the syntenic regions and mitochondrial gene organization are relatively conserved within the Vitaceae family compared to the other species examined. Finally, the phylogenetic tree was found to align closely with established taxonomic relationships among members of the rosid clade and underscores the close evolutionary relationship between Tetrastigma and Vitis. This study presents the first complete T. hemsleyanum mitogenome (809,173 bp, two circular chromosomes), revealing distinct codon usage, high repeat abundance (SSRs, tandem, dispersed), chloroplast-derived tRNAs, and conserved Vitaceae synteny. Key findings include C-to-U RNA editing, positive selection on ccmB, and phylogenetic confirmation of its close relationship to Vitis. This study elucidates mitochondrial genetics and provides a foundation for molecular breeding of Tetrastigma.