Characterization of chloroplast genome and phylogenetic analysis of the Elaeagnus hybrid variety E. 'viridis' (Elaeagnaceae).

Elaeagnus 'viridis', an artificial hybrid of E. macrophylla (♂) Thunb. (1784) × E. pungens (♀) Thunb. (1784), is known for its economic and ecological value. In this study, we sequenced and assembled the whole chloroplast (cp) genome of E. 'viridis'. The results showed that its cp genome was 152,284 bp long, showing a typical quadripartite structure and containing a large single-copy region (LSC, 82,299 bp), a small single-copy region (SSC, 18,239 bp), and a pair of inverted repeats (IRs, 51,746 bp). The cp genome contains 132 genes, including 86 protein-coding genes (PCGs), 38 tRNA genes, and 8 rRNA genes. Phylogenetic analysis based on 66 common PCGs revealed that E. 'viridis' is most closely related to its maternal parent E. pungens. The chloroplast genomic information reported in this study will shed some useful light for further genetic studies in the genus Elaeagnus.

Anthocyanin metabolism in Nelumbo: translational and post-translational regulation control transcription.

BACKGROUND: Lotus (Nelumbo Adans.) is used as an herbal medicine and the flowers are a source of natural flavonoids. 'Da Sajin', which was firstly found in the plateau area, is a natural mutant in flower color with red streamers dyeing around white petals. RESULTS: The LC-MS-MS results showed that eight anthocyanin compounds, including cyanidin 3-O-glucoside, cyanidin 3-O-galactoside, malvidin 3-O-galactoside, and malvidin 3-O-glucoside, were differentially enriched in red-pigmented tissues of the petals, whereas most of these metabolites were undetected in white tissues of the petals. Transcriptome profiling indicated that the relative high expression levels of structural genes, such as NnPAL, NnF3H, and NnANS, was inconsistent with the low anthocyanin concentration in white tissues. Members of the NnMYB and NnbHLH transcription factor families were presumed to play a role in the metabolic flux in the anthocyanin and proanthocyanidin biosynthetic pathway. The expression model of translational initiation factor, ribosomal proteins and SKP1-CUL1-F-box protein complex related genes suggested an important role for translational and post-translational network in anthocyanin biosynthesis. In addition, pathway analysis indicated that light reaction or photo destruction might be an important external cause for floral color determination in lotus. CONCLUSIONS: In this study, it is supposed that the natural lotus mutant 'Da Sajin' may have originated from a red-flowered ancestor. Partial loss of anthocyanin pigments in petals may result from metabolic disorder caused by light destruction. This disorder is mainly regulated at post translation and translation level, resulting in a non-inherited phenotype. These results contribute to an improved understanding of anthocyanin metabolism in lotus, and indicate that the translational and post-translational regulatory network determines the metabolic flux of anthocyanins and proanthocyanidins under specific environmental conditions.

Chromosome-Level Genome Assembly of a Fragrant Japonica Rice Cultivar 'Changxianggeng 1813' Provides Insights into Genomic Variations between Fragrant and Non-Fragrant Japonica Rice.

East Asia has an abundant resource of fragrant japonica rice that is gaining increasing interest among both consumers and producers. However, genomic resources and in particular complete genome sequences currently available for the breeding of fragrant japonica rice are still scarce. Here, integrating Nanopore long-read sequencing, Illumina short-read sequencing, and Hi-C methods, we presented a high-quality chromosome-level genome assembly (~378.78 Mb) for a new fragrant japonica cultivar 'Changxianggeng 1813', with 31,671 predicated protein-coding genes. Based on the annotated genome sequence, we demonstrated that it was the badh2-E2 type of deletion (a 7-bp deletion in the second exon) that caused fragrance in 'Changxianggeng 1813'. Comparative genomic analyses revealed that multiple gene families involved in the abiotic stress response were expanded in the 'Changxianggeng 1813' genome, which further supported the previous finding that no generalized loss of abiotic stress tolerance associated with the fragrance phenotype. Although the 'Changxianggeng 1813' genome showed high genomic synteny with the genome of the non-fragrant japonica rice cultivar Nipponbare, a total of 289,970 single nucleotide polymorphisms (SNPs), 96,093 small insertion-deletion polymorphisms (InDels), and 8690 large structure variants (SVs, >1000 bp) were identified between them. Together, these genomic resources will be valuable for elucidating the mechanisms underlying economically important traits and have wide-ranging implications for genomics-assisted breeding in fragrant japonica rice.

Characterization of the Glehnia littoralis Non-specific Phospholipase C Gene GlNPC3 and Its Involvement in the Salt Stress Response.

Glehnia littoralis is a medicinal halophyte that inhabits sandy beaches and has high ecological and commercial value. However, the molecular mechanism of salt adaptation in G. littoralis remains largely unknown. Here, we cloned and identified a non-specific phospholipase C gene (GlNPC3) from G. littoralis, which conferred lipid-mediated signaling during the salt stress response. The expression of GlNPC3 was induced continuously by salt treatment. Overexpression of GlNPC3 in Arabidopsis thaliana increased salt tolerance compared to wild-type (WT) plants. GlNPC3-overexpressing plants had longer roots and higher fresh and dry masses under the salt treatment. The GlNPC3 expression pattern revealed that the gene was expressed in most G. littoralis tissues, particularly in roots. The subcellular localization of GlNPC3 was mainly at the plasma membrane, and partially at the tonoplast. GlNPC3 hydrolyzed common membrane phospholipids, such as phosphotidylserine (PS), phosphoethanolamine (PE), and phosphocholine (PC). In vitro enzymatic assay showed salt-induced total non-specific phospholipase C (NPC) activation in A. thaliana GlNPC3-overexpressing plants. Plant lipid profiling showed a significant change in the membrane-lipid composition of A. thaliana GlNPC3-overexpressing plants compared to WT after the salt treatment. Furthermore, downregulation of GlNPC3 expression by virus-induced gene silencing in G. littoralis reduced the expression levels of some stress-related genes, such as SnRK2, P5SC5, TPC1, and SOS1. Together, these results indicated that GlNPC3 and GlNPC3-mediated membrane lipid change played a positive role in the response of G. littoralis to a saline environment.

Abundant Allelochemicals and the Inhibitory Mechanism of the Phenolic Acids in Water Dropwort for the Control of Microcystis aeruginosa Blooms.

In recent years, with the frequent global occurrence of harmful algal blooms, the use of plant allelopathy to control algal blooms has attracted special and wide attention. This study validates the possibility of turning water dropwort into a biological resource to inhibit the growth of harmful Microcystis aeruginosa blooms via allelopathy. The results revealed that there were 33 types of allelopathic compounds in the water dropwort culture water, of which 15 were phenolic acids. Regarding water dropwort itself, 18 phenolic acids were discovered in all the organs of water dropwort via a targeted metabolomics analysis; they were found to be mainly synthesized in the leaves and then transported to the roots and then ultimately released into culture water where they inhibited M. aeruginosa growth. Next, three types of phenolic acids synthesized in water dropwort, i.e., benzoic, salicylic, and ferulic acids, were selected to clarify their inhibitory effects on the growth of M. aeruginosa and their mechanism(s) of action. It was found that the inhibitory effect of phenolic acids on the growth of M. aeruginosa increased with the increase of the exposure concentration, although the algae cells were more sensitive to benzoic acid than to salicylic and ferulic acids. Further study indicated that the inhibitory effects of the three phenolic acids on the growth of M. aeruginosa were largely due to the simultaneous action of reducing the number of cells, damaging the integrity of the cell membrane, inhibiting chlorophyll a (Chl-a) synthesis, decreasing the values of F0 and Fv/Fm, and increasing the activity of the antioxidant enzymes (SOD, POD, and CAT) of M. aeruginosa. Thus, the results of this study indicate that both culture water including the rich allelochemicals in water dropwort and biological algae inhibitors made from water dropwort could be used to control the growth of noxious algae in the future.

Characterization and phylogenetic analysis of the complete chloroplast genome of Ilex rotunda, traditional Chinese medicine plant.

Ilex rotunda is a traditional Chinese medicine plant. In this study, we characterized the complete chloroplast (cp) genome sequence of I. rotunda to investigate its phylogenetic relationship. The cp genome of I. rotunda was 157,743 bp in length with 37.62% overall GC content, including a large single-copy (LSC) region of 87,060bp, a small single-copy (SSC) region of 18,432 bp, which were separated by a pair of inverted repeats (IRS) of 26,121 bp. The cp genome contained 133 genes, including 88 protein-coding genes, 37 tRNA genes, and 8 rRNA genes. Phylogenetic analysis based on whole cp genome sequences showed that I. rotunda is closely related to I. pubescens and I. polyneura.

The complete chloroplast genome sequence of Ilex chinensis Sims. (Aquifoliaceae), a folk herbal medicine plant in China.

The complete chloroplast (cp) genome of Ilex chinensis, an important economic plant with ornamental and ecological values, was sequenced to investigate its phylogenetic relationship. The entire cp genome of I. chinensis was 157,885 bp in length with 37.61% overall GC content, including a large single-copy (LSC) region of 87,289 bp, and a small single-copy (SSC) region of 18,388 bp, which were separated by a pair of inverted repeats (IRs) of 52,208 bp. The cp genome contained 135 genes, including 90 protein-coding genes, 37 tRNA genes, and eight rRNA genes. Phylogenetic analysis based on whole cp genome sequences showed that I. chinensis was closely related to I. szechwanensis and I. viridis species.

The complete chloroplast genome sequence and phylogenetic analysis of Ilex × Koehneana 'Wirt L. Winn' (Aquifoliaceae).

Ilex × Koehneana 'Wirt L. Winn', an important ornamental tree, has been widely distributed in southeastern China. In this study, we assembled and characterized the complete chloroplast (cp) genome of I. Koehneana to investigate its phylogenetic relationship. The whole cp genome of I. Koehneana is 157,538 bp, which contained a large single-copy (LSC) region of 87,055 bp and a small single-copy (SSC) region of 18,429 bp, and a pair of inverted repeats (IR) of 52,054 bp. A total of 137 genes, including 90 protein-coding genes, eight rRNAs, and 39 tRNAs, were identified. Phylogenetic analysis based on 74 conserved protein-coding genes revealed that I. Koehneana is closely related to I. 'tall boy'.

The complete chloroplast genome sequence and phylogenetic analysis of Ilex 'Beryl', a hybrid of Ilex cornuta × Ilex latifolia (Aquifoliaceae).

Ilex 'Beryl' is an ornamental and ecological tree widespread in southeastern China. In this study, the complete chloroplast (cp) genome of Ilex 'Beryl' was assembled and characterized to investigate its phylogenetic relationship. The entire cp genome of 'Beryl' was a typical quadripartite structure with 157,575 bp in length, including a large single-copy (LSC) region of 87,080 bp and a small single-copy (SSC) region of 18,427 bp, which were separated by a pair of inverted repeats (IRs) of 52,068 bp. There are 135 genes annotated, including 90 protein-coding genes, eight rRNA genes and 37 tRNA genes. Phylogenetic analysis based on whole cp genome sequences showed that 'Beryl' is closest to I. 'Emily Bruner' and I. 'tall boy'.

The complete chloroplast genome of Ilex 'Tall Boy', Ilex aquifolium × Ilex latifolia (Aquifoliaceae).

Thecomplete chloroplast (cp) genome of Ilex 'Tall Boy', an important economic plant with ornamental and ecological values, was sequenced to investigate its phylogenetic relationship. The entire cp genome of 'Tall Boy' was 157,527 bp in length with 37.65% overall GC content, including a large single-copy (LSC) region of 87,044 bp and a small single-copy (SSC) region of 18,429 bp, which were separated by a pair of inverted repeats (IRs) of 52,054 bp. The cp genome contained 135 genes, including 90 protein-coding genes, 37 tRNA genes, and 8 rRNA genes. Phylogenetic analysis based on whole cp genome sequences showed that 'Tall Boy' is closest to I. latifolia Thunb. species.

Selection and Validation of Reference Genes for Normalisation of Gene Expression in Glehnia littoralis.

Glehnia littoralis is an important medicinal halophyte-the dried root of which is used as Chinese herbal medicine. However, the use, selection and stability of reference genes are rarely verified in studies of G. littoralis, which hampers investigation of its salt tolerance and metabolism. In this study, we selected 13 candidate reference genes from the transcriptome data of G. littoralis-serine/threonine-protein phosphatase PP2A (PP2A), polyubiquitin 10 (UBQ10), actin (ACT), elongation factor 1-α (EF1-α), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), α-tubulin (α-TUB), ß-tubulin (ß-TUB), polypyrimidine tract-binding protein 1 (PTBP1), expressed protein 1 (EXP1), expressed protein 2 (EXP2), TIP41-like (TIP41), SAND family (SAND), and cyclophilin 2 (CYP2), and used qRT-PCR to analyse their expression levels in roots of G. littoralis treated with NaCl, polyethylene glycol (PEG), abscisic acid (ABA), and methyl jasmonate (MeJA), as well as in various organs of G. littoralis. The ΔCt, geNorm, NormFinder, and BestKeeper algorithms were used to assess the expression stability of the candidate reference genes and the results were then used to generate a comprehensive rank list with the RankAggreg R package. The most stable reference genes for normalisation were EXP1 and PP2A in response to NaCl, EXP2 and PP2A in response to ABA, CYP2 and α-TUB in response to MeJA, and ACT and EXP1 in the PEG and the organ subsets. GAPDH, ß-TUB, and UBQ10 exhibited low stability and so were unsuitable for normalisation. This study is the first systematic analysis of candidate reference genes in G. littoralis and will facilitate further investigation of normalisation of gene expression in G. littoralis.

The complete chloroplast genome of Ilex 'Emily Bruner', Ilex cornuta 'Burfordii' × Ilex latifolia (Aquifoliaceae).

Ilex 'Emily Bruner' is an important economic plant with ornamental and ecological functions in southeastern China. In this study, we characterized the complete chloroplast (cp) genome sequence of 'Emily Bruner' to investigate its phylogenetic relationship. The entire cp genome of 'Emily Bruner' was 157,216 bp in length with 37.68% overall GC content, including a large single-copy (LSC) region of 86,721 bp and a small single-copy (SSC) region of 18,427 bp, which were separated by a pair of inverted repeats (IRs) of 52,068 bp. The cp genome contained 135 genes, including 90 protein-coding genes, 37 tRNA genes, and 8 rRNA genes. Phylogenetic analysis based on whole cp genome sequences showed that 'Emily Bruner' is closest to I. cornuta species.

Nutrients resorption and stoichiometry characteristics of different-aged plantations of Larix kaempferi in the Qinling Mountains, central China.

Elucidating the stoichiometry and resorption patterns of multiple nutrients is of essential importance to holistically understanding plant nutrition and biogeochemical cycling. Although many studies on ecological stoichiometry have been carried out, surprisingly few of them were simultaneously done on the investigation of both nutrient resorption efficiency and stoichiometry for different-aged plantations of a perennial tree. Here, both green and senesced leaf samples were collected from four Larix kaempfer plantations aged of 8, 15, 22, and 32 years in the Qinling Mountains to examine nutrients resorption efficiency and stoichiometry characteristics. The results suggested that the nutrient concentrations of N, P, K, Ca, Mg, Al, and Fe in both green and senesced leaves of L. kaempferi (a deciduous conifer tree) did not show a regular change trends along the plantation ages in the Qinling Mountains. The concentrations of the most nutrients examined, except for Fe, in the green leaves were relatively lower than or close to the required physiological concentrations, suggesting a relative limitation of multiple nutrients exists in L. kaempferi for its above-ground biomass growth. The rank order of resorption efficiencies of four key nutrients (N, P, K, and Mg) was K (80.89%) > N (67.42%) > P (65.34%) > Mg (41.16%), whereas the nutrient Ca and Fe tended to accumulate in senesced leaves. Overall, the nutrient resorption efficiency of all examined elements did not exhibit a regular trend corresponding to the change of the plantation ages in L. kaempferi, but it was positively related to the nutrient concentrations in green leaves. The mean C:N and C:P ratios in the green and senesced leaves were significantly higher than those reported globally (on average). By contrast, the N:P ratio, at <14, was not only much lower than that reported for both China's flora and globally (on average), but it did suggest that the N nutrient limits growth of L. kaempferi in these plantations. Taken together, the results of this study are of substantial interest and value to forest managers and for the sustainable development of the Qinling forest ecosystems.

Gene Expression Profile in the Long-Living Lotus: Insights into the Heat Stress Response Mechanism.

Lotus (Nelumbo Adans) is an aquatic perennial plant that flourished during the middle Albian stage. In this study, we characterized the digital gene expression signatures for China Antique lotus under conditions of heat shock stress. Using RNA-seq technology, we sequenced four libraries, specifically, two biological replicates for control plant samples and two for heat stress samples. As a result, 6,528,866 to 8,771,183 clean reads were mapped to the reference genome, accounting for 92-96% total clean reads. A total of 396 significantly altered genes were detected across the genome, among which 315 were upregulated and 81 were downregulated by heat shock stress. Gene ontology (GO) enrichment of differentially expressed genes revealed protein folding, cell morphogenesis and cellular component morphogenesis as the top three functional terms under heat shock stress. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis led to the identification of protein processing in endoplasmic reticulum, plant-pathogen interactions, spliceosome, endocytosis, and protein export as significantly enriched pathways. Among the upregulated genes, small heat shock proteins (sHsps) and genes related to cell morphogenesis were particularly abundant under heat stress. Data from the current study provide valuable clues that may help elucidate the molecular events underlying heat stress response in China Antique lotus.

Molecular Evidence for the Hybrid Origin of Ilex dabieshanensis (Aquifoliaceae).

Ilex, the largest genus of dioecious woody plants, is a good study system to assess the role of hybridization in speciation and evolution. Ilex dabieshanensis, a tree endemic to Dabieshan Mountains region, was initially described as a new species. Based on morphological intermediacy and sympatric distribution with its putative parental species, I. cornuta and I. latifolia, we proposed it as a natural hybrid between them. In this study, we sequenced one chloroplast intergenic spacer (trnH-psbA) and two nuclear genes (gapC and nepGS) in I. dabieshanensis and its putative parental species to test the hybrid origin hypothesis. Our results showed that there were one to two differentially fixed sequence differences between I. cornuta and I. latifolia at the two nuclear genes. Twelve of the 14 individuals of I. dabieshanensis exhibited additivity in chromatograms on these differentially fixed sites at both nuclear genes, and the remaining two exhibited additivity in chromatograms on the fixed site at only the nepGS gene. Except one haplotype of I. cornuta at the nepGS gene, all of the haplotypes of I. cornuta at the two nuclear genes were well separated from those of I. latifolia, and most haplotypes of I. dabieshanensis were shared with those of I. cornuta and I. latifolia. Phylogenetic analysis of these haplotypes was largely consistent with haplotype network analysis. I. cornuta and I. latifolia differed by two nucleotide substitutions in the chloroplast intergenic spacer, and 12 individuals of I. dabieshanensis had the same sequences as I. latifolia, while the remaining two were identical with I. cornuta. The molecular data provide convincing evidence for the hybrid origin of I. dabieshanensis and asymmetrical direction of hybridization. One haplotype of I. cornuta at the nepGS gene was nested with those of I. latifolia, indicating introgression to I. cornuta.

[Patch diversity and spatial structure of wild Thymus quinquecostatus].

With the combination of ISSR (inter-simple sequence repeats), SRAP (sequence-related amplified polymorphism) and spatial autocorrelation, the genetic diversity and spatial structure per unit patch of three Huaiyuan populations of Thymus quinquecostatus in southeast China were analyzed. The results showed that there existed higher levels of genetic and clonal diversity among the patches within the wild T. quinquecostatus populations, with the percentage of polymorphic loc being 75.75%, Nei's gene diversity being 0.2537, Shannon's information index being 0.3811, percent of genetype (G/N) being 0.61, Simpson index (D) being 0.96, and Fager index (E) being 0.91. Analysis of molecular variance (AMOVA) showed that only 9.65% of genetic variation resided among the populations, while 90.35% of it resided among the individuals within the populations. No genotype patches in common were observed among the three populations. The spatial distribution of the same patches showed a concentrated distribution about 0-25 m, and that of different patches showed an inlaid distribution. Except for some locations that showed par correlations in the Huaiyuan populations of T. quinquecostatus, most locations lacked in spatial structure according to spatial autocorrelation analysis. The possible mechanism causing the establishment of the patches of T. quinquecostatus populations was due to seed dispersing, and the following clonal reproduction played important roles in patch development and population expanding.