Abstract
BACKGROUND: The confused taxonomic classification of Crucigenia is mainly inferred through morphological evidence and few nuclear genes and chloroplast genomic fragments. The phylogenetic status of C. quadrata, as the type species of Crucigenia, remains considerably controversial. Additionally, there are currently no reports on the chloroplast genome of Crucigenia. RESULTS: In this study, we utilize molecular phylogenetics and comparative genomics to show that C. quadrata belongs to Chlorophyceae rather than Trebouxiophyceae. The Bayesian and maximum likelihood (ML) phylogenetic trees support a monophyletic group of C. quadrata and Scenedesmaceae (Chlorophyceae) species. Our study presents the first complete chloroplast genome of C. quadrata, which is 197,184 bp in length and has a GC content of 31%. It has a typical quadripartite structure, and the chloroplast genome codons exhibit usage bias. Nucleotide diversity analysis highlights six genes (ccsA, psbF, chlN, cemA, rps3, rps18) as hotspots for genetic variation. Coding gene sequence divergence analyses indicate that four genes (cemA, clpP, psaA, rps3) are subject to positive selection. CONCLUSIONS: The determination of the phylogenetic status and the comparative chloroplast genomic analyses of C. quadrata will not only be useful in enhancing our understanding of the intricacy of Crucigenia taxonomy but also provide the important basis for studying the evolution of the incertae sedis taxa within Trebouxiophyceae.