Comparative plastome analysis of Blumea, with implications for genome evolution and phylogeny of Asteroideae

The genus Blumea (Asteroideae, Asteraceae) comprises about 100 species, including herbs, shrubs, and small trees. Previous studies have been unable to resolve taxonomic issues and the phylogeny of the genus Blumea due to the low polymorphism of molecular markers. Therefore, suitable polymorphic regions need to be identified. Here, we de novo assembled plastomes of the three Blumea species B. oxyodonta, B. tenella, and B. balsamifera and compared them with 25 other species of Asteroideae after correction of annotations. These species have quadripartite plastomes with similar gene content and genome organization comprising 113 genes, including 80 protein-coding, 29 transfer RNA, and 4 ribosomal RNA genes. The contraction and expansion of inverted repeats also show high similarities among the species. The comparative analysis of codon usage, amino acid frequency, microsatellite repeats, oligonucleotide repeats, and transition and transversion substitutions has revealed high resemblance among the newly assembled species of Blumea. We identified 10 highly polymorphic regions with nucleotide diversity above 0.02, including rps16-trnQ, ycf1, ndhF-rpl32, rps15, petN-psbM, and rpl32-trnL, and they may be suitable for the development of robust, authentic, and cost-effective markers for bar coding and inference of the phylogeny of the genus Blumea. Among these highly polymorphic regions, five regions also co-occurred with oligonucleotide repeats and support use of repeats as a proxy for the identification of polymorphic loci. The phylogenetic analysis revealed a close relationship between Blumea and Pluchea within the tribe Inuleae. Our study supports a sister relationship between “Astereae and Anthemideae,” while Gnaphalieae roots these two tribes, whereas in a previous study a sister relationship was reported between “Senecioneae and Anthemideae” and “Astereae and Gnaphalieae” using nuclear genome sequences. The conflicting phylogenetic signals observed at the tribal level between chloroplast and nuclear genome data require further investigation.Peer reviewe

Comparative Plastomics of Ashwagandha (Withania, Solanaceae) and Identification of Mutational Hotspots for Barcoding Medicinal Plants

Within the family Solanaceae, Withania is a small genus belonging to the Solanoideae subfamily. Here, we report the de novo assembled chloroplast genome sequences of W. coagulans, W. adpressa, and W. riebeckii. The length of these genomes ranged from 154,162 to 154,364 base pairs (bp). These genomes contained a pair of inverted repeats (IRa and IRb) ranging from 25,029 to 25,071 bp that were separated by a large single-copy (LSC) region of 85,635–85,765 bp and a small single-copy (SSC) region of 18,457–18,469 bp. We analyzed the structural organization, gene content and order, guanine-cytosine content, codon usage, RNA-editing sites, microsatellites, oligonucleotide and tandem repeats, and substitutions of Withania plastomes, which revealed high similarities among the species. Comparative analysis among the Withania species also highlighted 10 divergent hotspots that could potentially be used for molecular marker development, phylogenetic analysis, and species identification. Furthermore, our analyses showed that even three mutational hotspots (rps4-trnT, trnM-atpE, and rps15) were sufficient to discriminate the Withania species included in current study

The GASA Gene Family in Cacao (Theobroma cacao, Malvaceae): Genome Wide Identification and Expression Analysis

The gibberellic acid-stimulated Arabidopsis (GASA/GAST) gene family is widely distributed in plants and involved in various physiological and biological processes. These genes also provide resistance to abiotic and biotic stresses, including antimicrobial, antiviral, and antifungal. We are interested in characterizing the GASA gene family and determining its role in various physiological and biological process in Theobroma cacao. Here, we report 17 tcGASA genes distributed on six chromosomes in T. cacao. The gene structure, promoter region, protein structure and biochemical properties, expression, and phylogenetics of all tcGASAs were analyzed. Phylogenetic analyses divided tcGASA proteins into five groups. Among 17 tcGASA genes, nine segmentally duplicating genes were identified which formed four pairs and cluster together in phylogenetic tree. Differential expression analyses revealed that most of the tcGASA genes showed elevated expression in the seeds (cacao food), implying their role in seed development. The differential expression of tcGASAs was recorded between the tolerant and susceptible cultivars of cacao, which indicating their possible role as fungal resistant. Our findings provide new insight into the function, evolution, and regulatory system of the GASA family genes in T.cacao and may suggest new target genes for development of fungi-resistant cacao varieties in breeding programs

The GASA Gene Family in Cacao (Theobroma cacao, Malvaceae): Genome Wide Identification and Expression Analysis

The gibberellic acid-stimulated Arabidopsis (GASA/GAST) gene family is widely distributed in plants and involved in various physiological and biological processes. These genes also provide resistance to abiotic and biotic stresses, including antimicrobial, antiviral, and antifungal. We are interested in characterizing the GASA gene family and determining its role in various physiological and biological process in Theobroma cacao. Here, we report 17 tcGASA genes distributed on six chromosomes in T. cacao. The gene structure, promoter region, protein structure and biochemical properties, expression, and phylogenetics of all tcGASAs were analyzed. Phylogenetic analyses divided tcGASA proteins into five groups. Among 17 tcGASA genes, nine segmentally duplicating genes were identified which formed four pairs and cluster together in phylogenetic tree. Differential expression analyses revealed that most of the tcGASA genes showed elevated expression in the seeds (cacao food), implying their role in seed development. The differential expression of tcGASAs was recorded between the tolerant and susceptible cultivars of cacao, which indicating their possible role as fungal resistant. Our findings provide new insight into the function, evolution, and regulatory system of the GASA family genes in T.cacao and may suggest new target genes for development of fungi-resistant cacao varieties in breeding programs

Plastid genomics of Nicotiana (Solanaceae) : insights into molecular evolution, positive selection and the origin of the maternal genome of Aztec tobacco (Nicotiana rustica)

The genus Nicotiana of the family Solanaceae, commonly referred to as tobacco plants, are a group cultivated as garden ornamentals. Besides their use in the worldwide production of tobacco leaves, they are also used as evolutionary model systems due to their complex development history, which is tangled by polyploidy and hybridization. Here, we assembled the plastid genomes of five tobacco species, namely N. knightiana, N. rustica, N. paniculata, N. obtusifolia and N. glauca. De novo assembled tobacco plastid genomes showed typical quadripartite structure, consisting of a pair of inverted repeats (IR) regions (25,323–25,369 bp each) separated by a large single copy (LSC) region (86,510 –86,716 bp) and a small single copy (SSC) region (18,441–18,555 bp). Comparative analyses of Nicotiana plastid genomes showed similar GC content, gene content, codon usage, simple sequence repeats, oligonucleotide repeats, RNA editing sites and substitutions with currently available Solanaceae genomes sequences. We identified twenty highly polymorphic regions mostly belonging to intergenic spacer regions (IGS), which could be appropriate for the development of robust and cost-effective markers to infer the phylogeny of genus Nicotiana and family Solanaceae. Our comparative plastid genome analysis revealed that the maternal parent of the tetraploid N. rustica was the common ancestor of N. paniculata and N. knightiana, and the later species is more closely related to N. rustica. The relaxed molecular clock analyses estimated that the speciation event between N. rustica and N. knightiana appeared 0.56 Ma (HPD 0.65–0.46). The biogeographical analysis showed a south-to-north range expansion and diversification for N. rustica and related species, where N. undulata and N. paniculata evolved in North/Central Peru, while N. rustica developed in Southern Peru and separated from N. knightiana, which adapted to the Southern coastal climatic regimes. We further inspected selective pressure on protein-coding genes among tobacco species to determine if this adaptation process affected the evolution of plastid genes. These analyses indicated that four genes involved in different plastid functions, such as DNA replication (rpoA) and photosynthesis (atpB, ndhD and ndhF), came under positive selective pressure as a result of specific environmental conditions. Genetic mutations of the following genes might have contributed to the survival and better adaptation during the evolutionary history of tobacco species.Peer reviewe

Characterization of the Evolutionary Pressure on Anisodus tanguticus Maxim. with Complete Chloroplast Genome Sequence

Anisodus tanguticus Maxim. (Solanaceae), a traditional endangered Tibetan herb, is endemic to the Qinghai–Tibet Plateau. Here, we report the de novo assembled chloroplast (cp) genome sequences of A. tanguticus (155,765 bp). The cp contains a pair of inverted repeated (IRa and IRb) regions of 25,881 bp that are separated by a large single copy (LSC) region (86,516 bp) and a small single copy SSC (17,487 bp) region. A total of 132 functional genes were annotated in the cp genome, including 87 protein-coding genes, 37 tRNA genes, and 8 rRNA genes. Moreover, 199 simple sequence repeats (SSR) and 65 repeat structures were detected. Comparative plastome analyses revealed a conserved gene order and high similarity of protein-coding sequences. The A. tanguticus cp genome exhibits contraction and expansion, which differs from Przewalskia tangutica and other related Solanaceae species. We identified 30 highly polymorphic regions, mostly belonging to intergenic spacer regions (IGS), which may be suitable for the development of robust and cost-effective markers for inferring the phylogeny of the genus Anisodus and family Solanaceae. Analysis of the Ka/Ks ratios of the Hyoscyameae tribe revealed significant positive selection exerted on the cemA, rpoC2, and clpP genes, which suggests that protein metabolism may be an important strategy for A. tanguticus and other species in Hyoscyameae in adapting to the adverse environment on the Qinghai–Tibetan Plateau. Phylogenetic analysis revealed that A. tanguticus clustered closer with Hyoscyamus niger than P. tangutica. Our results provide reliable genetic information for future exploration of the taxonomy and phylogenetic evolution of the Hyoscyameae tribe and related species

Characterization of the Evolutionary Pressure on Anisodus tanguticus Maxim. with Complete Chloroplast Genome Sequence

Anisodus tanguticus Maxim. (Solanaceae), a traditional endangered Tibetan herb, is endemic to the Qinghai-Tibet Plateau. Here, we report the de novo assembled chloroplast (cp) genome sequences of A. tanguticus (155,765 bp). The cp contains a pair of inverted repeated (IRa and IRb) regions of 25,881 bp that are separated by a large single copy (LSC) region (86,516 bp) and a small single copy SSC (17,487 bp) region. A total of 132 functional genes were annotated in the cp genome, including 87 protein-coding genes, 37 tRNA genes, and 8 rRNA genes. Moreover, 199 simple sequence repeats (SSR) and 65 repeat structures were detected. Comparative plastome analyses revealed a conserved gene order and high similarity of protein-coding sequences. The A. tanguticus cp genome exhibits contraction and expansion, which differs from Przewalskia tangutica and other related Solanaceae species. We identified 30 highly polymorphic regions, mostly belonging to intergenic spacer regions (IGS), which may be suitable for the development of robust and cost-effective markers for inferring the phylogeny of the genus Anisodus and family Solanaceae. Analysis of the Ka/Ks ratios of the Hyoscyameae tribe revealed significant positive selection exerted on the cemA, rpoC2, and clpP genes, which suggests that protein metabolism may be an important strategy for A. tanguticus and other species in Hyoscyameae in adapting to the adverse environment on the Qinghai-Tibetan Plateau. Phylogenetic analysis revealed that A. tanguticus clustered closer with Hyoscyamus niger than P. tangutica. Our results provide reliable genetic information for future exploration of the taxonomy and phylogenetic evolution of the Hyoscyameae tribe and related species.Peer reviewe

Comparison of Chloroplast Genomes among Species of Unisexual and Bisexual Clades of the Monocot Family Araceae

The chloroplast genome provides insight into the evolution of plant species. We de novo assembled and annotated chloroplast genomes of four genera representing three subfamilies of Araceae: Lasia spinosa (Lasioideae), Stylochaeton bogneri, Zamioculcas zamiifolia (Zamioculcadoideae), and Orontium aquaticum (Orontioideae), and performed comparative genomics using these chloroplast genomes. The sizes of the chloroplast genomes ranged from 163,770 bp to 169,982 bp. These genomes comprise 113 unique genes, including 79 protein-coding, 4 rRNA, and 30 tRNA genes. Among these genes, 17–18 genes are duplicated in the inverted repeat (IR) regions, comprising 6–7 protein-coding (including trans-splicing gene rps12), 4 rRNA, and 7 tRNA genes. The total number of genes ranged between 130 and 131. The infA gene was found to be a pseudogene in all four genomes reported here. These genomes exhibited high similarities in codon usage, amino acid frequency, RNA editing sites, and microsatellites. The oligonucleotide repeats and junctions JSB (IRb/SSC) and JSA (SSC/IRa) were highly variable among the genomes. The patterns of IR contraction and expansion were shown to be homoplasious, and therefore unsuitable for phylogenetic analyses. Signatures of positive selection were seen in three genes in S. bogneri, including ycf2, clpP, and rpl36. This study is a valuable addition to the evolutionary history of chloroplast genome structure in Araceae

Complete Chloroplast Genomes of Anthurium huixtlense and Pothos scandens (Pothoideae, Araceae) : Unique Inverted Repeat Expansion and Contraction Affect Rate of Evolution

The subfamily Pothoideae belongs to the ecologically important plant family Araceae. Here, we report the chloroplast genomes of two species of the subfamily Pothoideae:Anthurium huixtlense(size: 163,116 bp) andPothos scandens(size: 164,719 bp). The chloroplast genome ofP. scandensshowed unique contraction and expansion of inverted repeats (IRs), thereby increasing the size of the large single-copy region (LSC: 102,956 bp) and decreasing the size of the small single-copy region (SSC: 6779 bp). This led to duplication of many single-copy genes due to transfer to IR regions from the small single-copy (SSC) region, whereas some duplicate genes became single copy due to transfer to large single-copy regions. The rate of evolution of protein-coding genes was affected by the contraction and expansion of IRs; we found higher mutation rates for genes that exist in single-copy regions as compared to those in IRs. We found a 2.3-fold increase of oligonucleotide repeats inP. scandenswhen compared withA. huixtlense, whereas amino acid frequency and codon usage revealed similarities. The ratio of transition to transversion mutations was 2.26 inP. scandensand 2.12 inA. huixtlense. Transversion mutations mostly translated in non-synonymous substitutions. The phylogenetic inference of the limited species showed the monophyly of the Araceae subfamilies. Our study provides insight into the molecular evolution of chloroplast genomes in the subfamily Pothoideae and family Araceae.Peer reviewe

Chloroplast genome evolution in the Dracunculus clade (Aroideae, Araceae)

Chloroplast (cp) genomes are considered important for the study of lineage-specific molecular evolution, population genetics, and phylogenetics. Our aim here was to elucidate the molecular evolution in cp genomes of species in the Dracunculus clade (Aroideae, Araceae). We report de novo assembled cp genomes for eight species from eight genera and also retrieved cp genomes of four species from the National Center for Biotechnology Information (NCBI). The cp genomes varied in size from 162,424 bp to 176,835 bp. Large Single Copy (LSC) region ranged in size from 87,141 bp to 95,475 bp; Small Single Copy (SSC) from 14,338 bp to 23,981 bp; and Inverted Repeats (IRa and IRb) from 25,131 bp to 32,708 bp. The expansion in inverted repeats led to duplication of ycf1 genes in four species. The genera showed high similarity in gene content and yielded 113 unique genes (79 protein-coding, 4 rRNA, and 30 tRNA genes). Codon usage, amino acid frequency, RNA editing sites, microsatellites repeats, transition and transversion substitutions, and synonymous and non-synonymous substitutions were also similar across the clade. A previous study reported deletion of ycf1, accD, psbE, trnL-CAA, and trnG-GCC genes in four Amorphophallus species. Our study supports conservative structure of cp genomes in the Dracunculus clade including Amorphophallus species and does not support gene deletion mentioned above. We also report suitable polymorphic loci based on comparative analyses of Dracunculus clade species, which could be useful for phylogenetic inference. Overall, the current study broad our knowledge about the molecular evolution of chloroplast genome in aroids.Peer reviewe