To systematically determine their phylogenetic relationships and develop molecular markers for species discrimination of Salvia bowleyana, S. splendens, and S. officinalis, we sequenced their chloroplast genomes using the Illumina Hiseq 2500 platform. The chloroplast genomes length of S. bowleyana, S. splendens, and S. officinalis were 151,387 bp, 150,604 bp, and 151,163 bp, respectively. The six genes ndhB, rpl2, rpl23, rps7, rps12, and ycf2 were present in the IR regions. The chloroplast genomes of S. bowleyana, S. splendens, and S. officinalis contain 29 tandem repeats; 35, 29, 24 simple-sequence repeats, and 47, 49, 40 interspersed repeats, respectively. The three specific intergenic sequences (IGS) of rps16-trnQ-UUG, trnL-UAA-trnF-GAA, and trnM-CAU-atpE were found to discriminate the 23 Salvia species. A total of 91 intergenic spacer sequences were identified through genetic distance analysis. The two specific IGS regions (trnG-GCC-trnM-CAU and ycf3-trnS-GGA) have the highest K2p value identified in the three studied Salvia species. Furthermore, the phylogenetic tree showed that the 23 Salvia species formed a monophyletic group. Two pairs of genus-specific DNA barcode primers were found. The results will provide a solid foundation to understand the phylogenetic classification of the three Salvia species. Moreover, the specific intergenic regions can provide the probability to discriminate the Salvia species between the phenotype and the distinction of gene fragments.

Background: Salvia bowleyana, S. splendens, and S. officinalis are globally distributed and have been widely used to treat coronary heart disease, liver tumors, and viral diseases. To systematically determine their phylogenetic relationship and develop molecular markers for species determination, we sequenced and assembled their chloroplast genomes, and analyzed the genome characteristics. Moreover, we compared the phylogenetic specification and divergence genes fragments of chloroplast genomes from the three Salvia species. Results: The length of S. bowleyana, S. splendens, and S. officinalis chloroplast genomes were 151387 bp, 150604 bp, and 151163 bp, respectively. The sizes of the large-single copy, small-single copy, and inverted repeat regions were 82772 bp, 17573 bp, and 51042 bp for S. bowleyana; 82181 bp, 17857 bp, and 50566 bp for S. splendens; 82429 bp, 17510 bp, and 51224 bp for S. officinalis, respectively. The GC contents of the three chloroplast genomes were 38.01%, 38.04%, 38.04%, partly. In the comparison of chloroplast genomes from Lamiaceae family, the six genes ndhB, rpl2, rpl23, rps7, rps12, and ycf2 were present in the IRs regions of all 41 chloroplast genomes in the Lamiaceae family. We found that one of the gene rpl20 was intact and stably occurred in all 41 species chloroplast genomes, however, another pseudogene one was lost in that of 40 species except the Dracocephalum heterophyllum. For the repeat analysis, 29 tandem repeats, 35, 29, 24 simple-sequence repeats(SSRs), and 47, 49, 40 interspersed repeats were identified in the three Salvia species chloroplast genomes based on the diverse requirements. The three specific intergenic sequences(IGS) of rps16-trnQ-UUG, trnL-UAA-trnF-GAA, and trnM-CAU-atpE were found to discriminate the certain species by comparing 23 Salvia chloroplast genomes. Six genes including rpl22, rps19, rpl2, ycf1, ndhF, and psbA were found in the highly diverse IR boundary regions. The genetic distance analysis of IGS showed the trnL-UAG-ccsA, rps16-trnQ-UUG, ccsA-ndhD, rps15-ycf1, and ndhE-ndhG regions had the higher variability. Furthermore, the phylogenetic tree inferred that the 23 Salvia species formed a monophyletic group. Lastly, two pairs of Genus-specific DNA barcode primers were identified, which can be used to amplify the part sequence of trnM-CAU-atpE and ccsA-ndhD region. Conclusions: We acquired the complete chloroplast genome of the three Salvia species, which will provide a solid foundation to understand their phylogenetic status in Salvia genus. Moreover, the research can provide the probability to discriminate the Salvia species compared with the genomics between the phenotype and the distinction of gene fragments .