Abstract
Macrotyloma is an underutilised legume genus within the tribe Phaseoleae (Fabaceae) that includes nutritionally and agronomically important crops such as horse gram (Macrotyloma uniflorum) and Kersting's groundnut (Macrotyloma geocarpum). Despite their importance, knowledge of the chloroplast (cp.) genome of this genus remains limited. In this study, we assembled and analysed the complete chloroplast genomes of three Macrotyloma species: M. uniflorum, M. geocarpum, and M. axillare. The chloroplast genomes were assembled into two isoforms that differ in the orientation of the small single-copy (SSC) region. Genome sizes ranged from 150,811 to 151,013 bp and exhibited the canonical quadripartite structure, comprising a pair of inverted repeats (IRa and IRb; 26,416-26,436 bp each), a large single-copy region (LSC; 80,229-80,446 bp), and a small single-copy region (SSC; 17,710-17,711 bp). Each genome encoded 110 unique genes, including 4 rRNA genes, 30 tRNA genes, and 76 protein-coding genes. All three species also possessed the ~ 50 kb inversion in the LSC region, a synapomorphy shared among a large clade within the Papilionoideae subfamily of Fabaceae. Although overall chloroplast genome structure and organisation were highly conserved among Macrotyloma species, gene-wise nucleotide diversity analysis identified seven relatively variable genes: rps18, rps15, ccsA, ndhA, ycf1, ycf4, and psaI. Phylogenomic analysis based on complete chloroplast genomes robustly resolved Macrotyloma as a monophyletic group within the Phaseolinae clade of the Papilionoideae subfamily. Within the genus, M. uniflorum and M. axillare formed a strongly supported sister pair, with M. geocarpum sister to this clade. Overall, this study provides valuable insights into chloroplast genome evolution in Macrotyloma and enhances understanding of its phylogenetic placement within Phaseoleae, offering genomic resources for future evolutionary, taxonomic, and conservation studies of this underutilised legume genus.