In this study, we assembled and annotated the chloroplast (cp) genome of the Euonymus species Euonymus fortunei, Euonymus phellomanus, and Euonymus maackii, and performed a series of analyses to investigate gene structure, GC content, sequence alignment, and nucleic acid diversity, with the objectives of identifying positive selection genes and understanding evolutionary relationships. The results indicated that the Euonymus cp genome was 156,860–157,611bp in length and exhibited a typical circular tetrad structure. Similar to the majority of angiosperm chloroplast genomes, the results yielded a large single-copy region (LSC) (85,826–86,299bp) and a small single-copy region (SSC) (18,319–18,536bp), separated by a pair of sequences (IRA and IRB; 26,341–26,700bp) with the same encoding but in opposite directions. The chloroplast genome was annotated to 130–131 genes, including 85–86 protein coding genes, 37 tRNA genes, and eight rRNA genes, with GC contents of 37.26–37.31%. The GC content was variable among regions and was highest in the inverted repeat (IR) region. The IR boundary of Euonymus happened expanding resulting that the rps19 entered into IR region and doubled completely. Such fluctuations at the border positions might be helpful in determining evolutionary relationships among Euonymus. The simple-sequence repeats (SSRs) of Euonymus species were composed primarily of single nucleotides (A)n and (T)n, and were mostly 10–12bp in length, with an obvious A/T bias. We identified several loci with suitable polymorphism with the potential use as molecular markers for inferring the phylogeny within the genus Euonymus. Signatures of positive selection were seen in rpoB protein encoding genes. Based on data from the whole chloroplast genome, common single copy genes, and the LSC, SSC, and IR regions, we constructed an evolutionary tree of Euonymus and related species, the results of which were consistent with traditional taxonomic classifications. It showed that E. fortunei sister to the Euonymus japonicus, whereby E. maackii appeared as sister to Euonymus hamiltonianus. Our study provides important genetic information to support further investigations into the phylogenetic development and adaptive evolution of Euonymus species.

Pyrus hopeiensis is a valuable wild resource of Pyrus in the Rosaceae. Due to its limited distribution and population decline, it has been listed as one of the “wild plants with a tiny population” in China. To date, few studies have been conducted on P. hopeiensis. This paper offers a systematic review of P. hopeiensis, providing a basis for the conservation and restoration of P. hopeiensis resources. In this study, the chloroplast genomes of two different genotypes of P. hopeiensis, P. ussuriensis Maxin. cv. Jingbaili, P. communis L. cv. Early Red Comice, and P. betulifolia were sequenced, compared and analyzed. The two P. hopeiensis genotypes showed a typical tetrad chloroplast genome, including a pair of inverted repeats encoding the same but opposite direction sequences, a large single copy (LSC) region, and a small single copy (SSC) region. The length of the chloroplast genome of P. hopeiensis HB-1 was 159,935 bp, 46 bp longer than that of the chloroplast genome of P. hopeiensis HB-2. The lengths of the SSC and IR regions of the two Pyrus genotypes were identical, with the only difference present in the LSC region. The GC content was only 0.02% higher in P. hopeiensis HB-1. The structure and size of the chloroplast genome, the gene species, gene number, and GC content of P. hopeiensis were similar to those of the other three Pyrus species. The IR boundary of the two genotypes of P. hopeiensis showed a similar degree of expansion. To determine the evolutionary history of P. hopeiensis within the genus Pyrus and the Rosaceae, 57 common protein-coding genes from 36 Rosaceae species were analyzed. The phylogenetic tree showed a close relationship between the genera Pyrus and Malus, and the relationship between P. hopeiensis HB-1 and P. hopeiensis HB-2 was the closest.

Pyrus hopeiensis is a valuable but endangered wild resource in the genus Pyrus. It has been listed as one of the 120 wild species with tiny population in China. The specie has been little studied. A preliminary study of propagation modes in P. hopeiensis was performed through seed propagation, hybridization, self-crossing trials, bud grafting, branch grafting, and investigations of natural growth. The results showed that the population size of P. hopeiensis was very small, the distribution range was limited, and the habitat was extremely degraded. In the wild population, natural hybridization and root tiller production were the major modes of propagation. Whole genome re-sequencing of the 23 wild and cultivated accessions from Pyrus species collected was performed using an Illumina HiSeq sequencing platform. The sequencing depth range was 26.56x−44.85x and the average sequencing depth was 32x. Phylogenetic tree and principal component analyses (PCA) based on SNPs showed that the wild Pyrus species, such as PWH06, PWH07, PWH09, PWH10, PWH13, and PWH17, were closely related to both P. hopeiensis HB-1 and P. hopeiensis HB-2. Using these results in combination with morphological characteristics, it speculated that P. hopeiensis populations may form a natural hybrid group with frequent gene exchanges between and within groups. A selective elimination analysis on the P. hopeiensis population were performed using Fst and π radio and a total of 381 overlapping genes including SAUR72, IAA20, HSFA2, and RKP genes were obtained. These genes were analyzed by gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) function enrichment. And four KEGG pathways, including lysine degradation, sphingolipid metabolism, other glycan degradation, and betaine biosynthesis were significantly enriched in the P. hopeiensis population. Our study provided information on genetic variation, evolutionary relationships, and gene enrichment in P. hopeiensis population. These data will help reveal the evolutionary history and origin of P. hopeiensis and provide guidelines for subsequent research on the locations of functional genes.

In this study, we assembled and annotated the chloroplast (cp) genomes of four Ligustrum species, L. sinense, L. obtusifolium, L. vicaryi, and L. ovalifolium ‘Aureum’. Including six other published Ligustrum species, we compared various characteristics such as gene structure, sequence alignment, codon preference, and nucleic acid diversity, and performed positive-selection genes screening and phylogenetic analysis. The results showed that the cp genome of Ligustrum was 162,185–166,800 bp in length, with a circular tetrad structure, including a large single-copy region (86,885–90,106 bp), a small single-copy region (11,446–11,499 bp), and a pair of IRa and IRb sequences with the same coding but in opposite directions (31,608–32,624 bp). This structure is similar to the cp genomes of most angiosperms. We found 132–137 genes in the cp genome of Ligustrum, including 89–90 protein-coding genes, 35–39 tRNAs, and 8 rRNAs. The GC content was 37.93–38.06% and varied among regions, with the IR region having the highest content. The single-nucleotide (A/T)n was dominant in simple-sequence repeats of the Ligustrum cp genome, with an obvious A/T preference. Six hotspot regions were identified from multiple sequence alignment of Ligustrum; the ycf1 gene region and the clpP1 exon region can be used as potential DNA barcodes for the identification and phylogeny of the genus Ligustrum. Branch-site model and Bayes empirical Bayes (BEB) analysis showed that four protein-coding genes (accD, clpP, ycf1, and ycf2) were positively selected, and BEB analysis showed that accD and rpl20 had positively selected sites. A phylogenetic tree of Oleaceae species was constructed based on the whole cp genomes, and the results were consistent with the traditional taxonomic results. The phylogenetic results showed that genus Ligustrum is most closely related to genus Syringa. Our study provides important genetic information to support further investigations of the phylogenetic development and adaptive evolution of Ligustrum species.

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