The order Nymphaeales, consisting of three families with a record of eight genera, has gained significant interest from botanists, probably due to its position as a basal angiosperm. The phylogenetic relationships within the order have been well studied; however, a few controversial nodes still remain in the Nymphaeaceae. The position of the Nuphar genus and the monophyly of the Nymphaeaceae family remain uncertain. This study adds to the increasing number of the completely sequenced plastid genomes of the Nymphaeales and applies a large chloroplast gene data set in reconstructing the intergeneric relationships within the Nymphaeaceae. Five complete chloroplast genomes were newly generated, including a first for the monotypic Euryale genus. Using a set of 66 protein-coding genes from the chloroplast genomes of 17 taxa, the phylogenetic position of Nuphar was determined and a monophyletic Nymphaeaceae family was obtained with convincing statistical support from both partitioned and unpartitioned data schemes. Although genomic comparative analyses revealed a high degree of synteny among the chloroplast genomes of the ancient angiosperms, key minor variations were evident, particularly in the contraction/expansion of the inverted-repeat regions and in RNA-editing events. Genome structure, and gene content and arrangement were highly conserved among the chloroplast genomes. The intergeneric relationships defined in this study are congruent with those inferred using morphological data.

Many plant species exhibit different leaf morphologies within a single plant, or heterophylly. The molecular mechanisms regulating this phenomenon, however, have remained elusive. In this study, the transcriptomes of submerged and floating leaves of an aquatic heterophyllous plant, Potamogeton octandrus Poir, at different stages of development, were sequenced using high-throughput sequencing (RNA-Seq), in order to aid gene discovery and functional studies of genes involved in heterophylly. A total of 81,103 unigenes were identified in submerged and floating leaves and 6,822 differentially expressed genes (DEGs) were identified by comparing samples at differing time points of development. KEGG pathway enrichment analysis categorized these unigenes into 128 pathways. A total of 24,025 differentially expressed genes were involved in carbon metabolic pathways, biosynthesis of amino acids, ribosomal processes, and plant-pathogen interactions. In particular, KEGG pathway enrichment analysis categorized a total of 70 DEGs into plant hormone signal transduction pathways. The high-throughput transcriptomic results presented here highlight the potential for understanding the molecular mechanisms underlying heterophylly, which is still poorly understood. Further, these data provide a framework to better understand heterophyllous leaf development in P. octandrus via targeted studies utilizing gene cloning and functional analyses.

The order Nymphaeales, consisting of three families with a record of eight genera, has gained significant interest from botanists probably due to its position as a basal-angiosperm. The phylogenetic relationships within the order have well been studied and resolved; however, a few controversial nodes still remain in the Nymphaeaceae including the position of the genus Nuphar. The position of the genus Nuphar and the monophyly of the Nymphaeaceae family remain uncertain. This study adds to the increasing number of completely sequenced plastid genomes of the Nymphaeales and applies large chloroplast gene data set in reconstructing the intergeneric relationships within the Nymphaeaceae. Five complete chloroplast genomes were newly generated, including a first one for the monotypic genus Euryale. Using a set of 66 protein coding genes from the chloroplast genomes of 17 taxa, the phylogenetic position of Nuphar was determined and a monophyletic Nymphaeaceae family was obtained with a convincing statistical support from both partitioned and unpartitioned data schemes. Although genomic comparative analyses revealed a high degree of synteny among the chloroplast genomes of the ancient angiosperms, key minor variations were evident particularly in the contraction/expansion of the Inverted Repeat regions and in RNA editing events. Genome structure, gene content and arrangement were highly conserved among the chloroplast genomes.

Members of the aquatic plant genus Aponogeton are widely used commercially in aquariums because of their variable leaf shape and unique inflorescences. However, due to extensive similarity between species in this genus, morphological characters are generally inadequate for taxonomic classification. Currently, molecular makers available for taxonomic and phylogenetic studies of Aponogeton are limited. One approach to clarifying relationships between species in these complex groups is to use divergence hotspot regions within the genome. Here, we sequenced and analyzed the plastomes of five Aponogeton species collected from China, Zambia, and Kenya, and subsequently screened these plastomes for divergent DNA hotspots. The five plastomes are circular structures with sizes ranging from 154,167 bp to 154,860 bp. The Large and the Small Single Copies are separated by two Inverted Repeats. One hundred and thirteen unique genes were identified including 79 protein-coding, 30 tRNA, and four rRNA genes. We found that the most abundant repeats in all but one species were mononucleotide repeats (A/T) and that there were 23 potential RNA ending sites. Interestingly, a ~3 kb inversion, which includes the accD gene, was detected within the Asian species of Aponogeton . The inversion may be related to more frequent exchanges between this region and the nuclear genome. Furthermore, we detected mutational hotspot sites among the five Aponogeton species. Three of these hotspots are intergenic spacer regions ( accD-psaI , rbcL-accD and trnH-GUG-psbA ) that might be suitable for use as barcodes to resolve intra-generic relationships. We also identified four highly variable protein-coding genes ( ccsA , rpl22 , rps16 and ycf1 ) may be used as barcodes to resolve the higher-level phylogenies. Our study will provide valuable molecular resources for the taxonomic and phylogenomic study of the complex genus Aponogeton .

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Many plant species exhibit heterophylly, displaying different leaves upon a single plant.The molecular mechanisms regulating this phenomenon, however, have remained elusive.In this study, the transcriptomes of submerged and floating leaves of an aquatic heterophyllous plant, Potamogeton octandrus Poir, were sequenced using a highthroughput sequencing technique (RNA-Seq), which aims to assist with the gene discovery and functional studies of genes involved in heterophyllous leaf development. A total of 81,103 unigenes were identified from the submerged and floating leaves, and a total of 6,822 differentially expressed genes (DEGs) were identified by comparing the samples from each developmental stage. KEGG pathway enrichment analysis categorized these unigenes into 128 pathways (p-value < 10 -5 ). A total of 24,025 differentially expressed genes were involved in the carbon metabolic pathway, biosynthesis of amino acids, ribosomes, and plant-pathogen interaction. KEGG pathway enrichment analysis categorized a total of 70 DEGs into plant hormone signal transduction pathways. This study describes the initial results of the high-throughput transcriptome sequencing of heterophylly. Understanding the transcriptomes of floating and submerged leaves of the aquatic plant P. octandrus will assist with gene cloning and functional studies of genes involved in leaf development. This is especially the case with those involved in heterophyllous leaf development. 36 Understanding the transcriptomes of floating and submerged leaves of the aquatic plant P.37 octandrus will assist with gene cloning and functional studies of genes involved in leaf 38 development. This is especially the case with those involved in heterophyllous leaf development. (Minorsky, 2003; Zotz, Wilhelm & Becker, 45 2011). In some cases, heterophylly is believed to be an adaptive response to the environment, 46 and it has been linked to an increase in fitness (Cook & Johnson, 1968; Wells & Pigliucci, 2000; 47 Minorsky, 2003). For example, the heterophylly of aquatic plants may increase their fitness by 48 decreasing leaf damage, decreasing water loss, enhancing photosynthesis, or promoting sexual 49 reproductive success (Winn, 1999a,b; Wells & Pigliucci, 2000; Minorsky, 2003; Zhang et al., 50 2009;Zotz, Wilhelm & Becker, 2011). Accordingly, heterophylly has been used as a model 51 system for studying gene-environment interactions (Pigliucci, 2010; Nakayama et al., 2014). 52In the past century, numerous studies have been conducted to describe morphological 53 changes in heterophyllous plants in response to environmental factors such as CO 2 concentration, 54 oxygen capacity, salt concentration, temperature, water level, seasonal change, and light intensity 55 and quality (McCallum, 1902;Arber, 1920; Fassett, 1930; Sculthorpe, 1967; Cook & Johnson, 56 1968;Bodkin, Spence & Weeks, 1980; Deschamp & Cooke, 1984; Titus & Sullivan, 2001 88 on a single plant is controlled through multiple signalling pathways (Lin & Yang, 1999; Hsu et 89 al., 2001). Thus, wh...

Many plant species exhibit heterophylly, displaying different leaves upon a single plant.The molecular mechanisms regulating this phenomenon, however, have remained elusive.In this study, the transcriptomes of submerged and floating leaves of an aquatic heterophyllous plant, Potamogeton octandrus Poir, were sequenced using a highthroughput sequencing technique (RNA-Seq), which aims to assist with the gene discovery and functional studies of genes involved in heterophyllous leaf development. A total of 81,103 unigenes were identified from the submerged and floating leaves, and a total of 6,822 differentially expressed genes (DEGs) were identified by comparing the samples from each developmental stage. KEGG pathway enrichment analysis categorized these unigenes into 128 pathways (p-value < 10 -5 ). A total of 24,025 differentially expressed genes were involved in the carbon metabolic pathway, biosynthesis of amino acids, ribosomes, and plant-pathogen interaction. KEGG pathway enrichment analysis categorized a total of 70 DEGs into plant hormone signal transduction pathways. This study describes the initial results of the high-throughput transcriptome sequencing of heterophylly. Understanding the transcriptomes of floating and submerged leaves of the aquatic plant P. octandrus will assist with gene cloning and functional studies of genes involved in leaf development. This is especially the case with those involved in heterophyllous leaf development. 26 Potamogeton octandrus Poir, were sequenced using a high-throughput sequencing technique 27 (RNA-Seq), which aims to assist with the gene discovery and functional studies of genes 28 involved in heterophyllous leaf development. A total of 81,103 unigenes were identified from the 29 submerged and floating leaves, and a total of 6,822 differentially expressed genes (DEGs) were 30 identified by comparing the samples from each developmental stage. KEGG pathway enrichment 31 analysis categorized these unigenes into 128 pathways (p-value < 10-5 ). A total of 24,025 32 differentially expressed genes were involved in the carbon metabolic pathway, biosynthesis of 33 amino acids, ribosomes, and plant-pathogen interaction. KEGG pathway enrichment analysis 34 categorized a total of 70 DEGs into plant hormone signal transduction pathways. This study 35 describes the initial results of the high-throughput transcriptome sequencing of heterophylly. PeerJ Preprints36 Understanding the transcriptomes of floating and submerged leaves of the aquatic plant P.37 octandrus will assist with gene cloning and functional studies of genes involved in leaf 38 development. This is especially the case with those involved in heterophyllous leaf development. (Minorsky, 2003; Zotz, Wilhelm & Becker, 45 2011). In some cases, heterophylly is believed to be an adaptive response to the environment, 46 and it has been linked to an increase in fitness (Cook & Johnson, 1968; Wells & Pigliucci, 2000; 47 Minorsky, 2003). For example, the heterophylly of aquatic plants may increase their fitness by 48 decr...