Sequence characteristics, genetic diversity and phylogenetic analysis of the Cucurbita ficifolia (Cucurbitaceae) chloroplasts genome.

BACKGROUND: Curcubita ficifolia Bouché (Cucurbitaceae) has high value as a food crop and medicinal plant, and also has horticultural value as rootstock for other melon species. China is home to many different cultivars, but the genetic diversity of these resources and the evolutionary relationships among them, as well as the differences between C. ficifolia and other Cucurbita species, remain unclear. RESULTS: We investigated the chloroplast (cp) genomes of 160 C. ficifolia individuals from 31 populations in Yunnan, a major C. ficifolia production area in China. We found that the cp genome of C. ficifolia is ~151 kb and contains 128 genes, of which 86 are protein coding genes, 34 encode tRNA, and eight encode rRNAs. We also identified 64 SSRs, mainly AT repeats. The cp genome was found to contain a total of 204 SNP and 57 indels, and a total of 21 haplotypes were found in the 160 study individuals. The reverse repeat (IR) region of C. ficifolia contained a few differences compared with this region in the six other Cucurbita species. Sequence difference analysis demonstrated that most of the variable regions were concentrated in the single copy (SC) region. Moreover, the sequences of the coding regions were found to be more similar among species than those of the non-coding regions. The phylogenies reconstructed from the cp genomes of 61 representative species of Cucurbitaceae reflected the currently accepted classification, in which C. ficifolia is sister to the other Cucurbita species, however, different interspecific relationships were found between Cucurbita species. CONCLUSIONS: These results will be valuable in the classification of C. ficifolia genetic resources and will contribute to our understanding of evolutionary relationships within the genus Cucurbita.

Integrated metabolome and transcriptome analyses reveal the molecular mechanism underlying dynamic metabolic processes during taproot development of Panax notoginseng.

BACKGROUND: Panax notoginseng (Burk) F. H. Chen is one of the most famous Chinese traditional medicinal plants. The taproot is the main organ producing triterpenoid saponins, and its development is directly linked to the quality and yield of the harvested P. notoginseng. However, the mechanisms underlying the dynamic metabolic changes occurring during taproot development of P. notoginseng are unknown. RESULTS: We carried out metabolomic and transcriptomic analyses to investigate metabolites and gene expression during the development of P. notoginseng taproots. The differentially accumulated metabolites included amino acids and derivatives, nucleotides and derivatives, and lipids in 1-year-old taproots, flavonoids and terpenoids in 2- and 3-year-old taproots, and phenolic acids in 3-year-old taproots. The differentially expressed genes (DEGs) are related to phenylpropanoid biosynthesis, metabolic pathway and biosynthesis of secondary metabolites at all three developmental stages. Integrative analysis revealed that the phenylpropanoid biosynthesis pathway was involved in not only the development of but also metabolic changes in P. notoginseng taproots. Moreover, significant accumulation of triterpenoid saponins in 2- and 3-year-old taproots was highly correlated with the up-regulated expression of cytochrome P450s and uridine diphosphate-dependent glycosyltransferases genes. Additionally, a gene encoding RNase-like major storage protein was identified to play a dual role in the development of P. notoginseng taproots and their triterpenoid saponins synthesis. CONCLUSIONS: These results elucidate the molecular mechanism underlying the accumulation of and change relationship between primary and secondary metabolites in P. notoginseng taproots, and provide a basis for the quality control and genetic improvement of P. notoginseng.

Genome-wide identification and molecular characterization of CRK gene family in cucumber (Cucumis sativus L.) under cold stress and sclerotium rolfsii infection.

BACKGROUND: The plant cysteine-rich receptor-like kinases (CRKs) are a large family having multiple roles, including defense responses under both biotic and abiotic stress. However, the CRK family in cucumbers (Cucumis sativus L.) has been explored to a limited extent. In this study, a genome-wide characterization of the CRK family has been performed to investigate the structural and functional attributes of the cucumber CRKs under cold and fungal pathogen stress. RESULTS: A total of 15 C. sativus CRKs (CsCRKs) have been characterized in the cucumber genome. Chromosome mapping of the CsCRKs revealed that 15 genes are distributed in cucumber chromosomes. Additionally, the gene duplication analysis of the CsCRKs yielded information on their divergence and expansion in cucumbers. Phylogenetic analysis divided the CsCRKs into two clades along with other plant CRKs. Functional predictions of the CsCRKs suggested their role in signaling and defense response in cucumbers. The expression analysis of the CsCRKs by using transcriptome data and via qRT-PCR indicated their involvement in both biotic and abiotic stress responses. Under the cucumber neck rot pathogen, Sclerotium rolfsii infection, multiple CsCRKs exhibited induced expressions at early, late, and both stages. Finally, the protein interaction network prediction results identified some key possible interacting partners of the CsCRKs in regulating cucumber physiological processes. CONCLUSIONS: The results of this study identified and characterized the CRK gene family in cucumbers. Functional predictions and validation via expression analysis confirmed the involvement of the CsCRKs in cucumber defense response, especially against S. rolfsii. Moreover, current findings provide better insights into the cucumber CRKs and their involvement in defense responses.

The effect of environmental factors on the genetic differentiation of Cucurbita ficifolia populations based on whole-genome resequencing.

BACKGROUND: Cucurbita ficifolia is one of the squash species most resistant to fungal pathogens, and has especially high resistance to melon Fusarium wilt. This species is therefore an important germplasm resource for the breeding of squash and melon cultivars. RESULTS: Whole-genome resequencing of 223 individuals from 32 populations in Yunnan Province, the main cucurbit production area in China, was performed and 3,855,120 single-nucleotide polymorphisms (SNPs) and 1,361,000 InDels were obtained. SNP analysis suggested that levels of genetic diversity in C. ficifolia were high, but that different populations showed no significant genetic differentiation or geographical structure, and that individual C. ficifolia plants with fruit rinds of a similar color did not form independent clusters. A Mantel test conducted in combination with geographical distance and environmental factors suggested that genetic distance was not correlated with geographical distance, but had a significant correlation with environmental distance. Further associations between the genetic data and five environmental factors were analyzed using whole-genome association analysis. SNPs associated with each environmental factor were investigated and genes 250 kb upstream and downstream from associated SNPs were annotated. Overall, 15 marker-trait-associated SNPs (MTAs) and 293 genes under environmental selection were identified. The identified genes were involved in cell membrane lipid metabolism, macromolecular complexes, catalytic activity and other related aspects. Ecological niche modeling was used to simulate the distribution of C. ficifolia across time, from the present and into the future. We found that the area suitable for C. ficifolia changed with the changing climate in different periods. CONCLUSIONS: Resequencing of the C. ficifolia accessions has allowed identification of genetic markers, such as SNPs and InDels. The SNPs identified in this study suggest that environmental factors mediated the formation of the population structure of C. ficifolia in China. These SNPs and Indels might also contribute to the variation in important pathways of genes for important agronomic traits such as yield, disease resistance and stress tolerance. Moreover, the genome resequencing data and the genetic markers identified from 223 accessions provide insight into the genetic variation of the C. ficifolia germplasm and will facilitate a broad range of genetic studies.

Transcriptomic and proteomic analyses of Cucurbita ficifolia Bouché (Cucurbitaceae) response to Fusarium oxysporum f.sp. cucumerium.

BACKGROUND: Fusarium oxysporum f. sp. cucumerinum (FOC) is the causal agent of cucumber Fusarium wilt, which can cause extensive damages and productivity losses. Cucurbita ficifolia Bouché (Cucurbitaceae) is usually used as rootstock for cucumber because of its excellent resistance to Fusarium wilt. Our previous study found that C.ficifolia has high FOC resistance, the underlying mechanism of which is unclear. RESULTS: Transcriptome and proteome profiling was performed on the basis of RNA-Seq and isobaric tag for relative and absolute quantitation technology to explore the molecular mechanisms of the response of Cucurbita ficifolia Bouché to Fusarium oxysporum f. sp. cucumerium infection. Comparative analyses revealed that 1850 genes and 356 protein species were differentially regulated at 2d and 4d after FOC inoculation. However, correlation analysis revealed that only 11 and 39 genes were differentially regulated at both the transcriptome and proteome levels after FOC inoculation at 2d and 4d, respectively. After FOC inoculation, plant hormones signal transduction, transcription factors were stimulated, whereas wax biosynthesis and photosynthesis were suppressed. Increased synthesis of oxidative-redox proteins is involved in resistance to FOC. CONCLUSIONS: This study is the first to reveal the response of C. ficifolia leaf to FOC infection at the transcriptome and proteome levels, and to show that FOC infection activates plant hormone signaling and transcription factors while suppressing wax biosynthesis and photosynthesis. The accumulation of oxidative-redox proteins also plays an important role in the resistance of C. ficifolia to FOC. Results provide new information regarding the processes of C. ficifolia leaf resistance to FOC and will contribute to the breeding of cucumber rootstock with FOC resistance.

Identification of Potential Dipeptidyl Peptidase (DPP)-IV Inhibitors among Moringa oleifera Phytochemicals by Virtual Screening, Molecular Docking Analysis, ADME/T-Based Prediction, and In Vitro Analyses.

Moringa oleifera Lam. (MO) is called the "Miracle Tree" because of its extensive pharmacological activity. In addition to being an important food, it has also been used for a long time in traditional medicine in Asia for the treatment of chronic diseases such as diabetes and obesity. In this study, by constructing a library of MO phytochemical structures and using Discovery Studio software, compounds were subjected to virtual screening and molecular docking experiments related to their inhibition of dipeptidyl peptidase (DPP-IV), an important target for the treatment of type 2 diabetes. After the four-step screening process, involving screening for drug-like compounds, predicting the absorption, distribution, metabolism, excretion, and toxicity (ADME/T) of pharmacokinetic properties, LibDock heatmap matching analysis, and CDOCKER molecular docking analysis, three MO components that were candidate DPP-IV inhibitors were identified and their docking modes were analyzed. In vitro activity verification showed that all three MO components had certain DPP-IV inhibitory activities, of which O-Ethyl-4-[(α-l-rhamnosyloxy)-benzyl] carbamate (compound 1) had the highest activity (half-maximal inhibitory concentration [IC50] = 798 nM). This study provides a reference for exploring the molecular mechanisms underlying the anti-diabetic activity of MO. The obtained DPP-IV inhibitors could be used for structural optimization and in-depth in vivo evaluation.

Population structure of wild soybean (Glycine soja) based on SLAF-seq have implications for its conservation.

Background: Glycine soja Sieb. & Zucc. is the wild ancestor from which the important crop plant soybean was bred. G. soja provides important germplasm resources for the breeding and improvement of cultivated soybean crops, however the species is threatened by habitat loss and fragmentation, and is experiencing population declines across its natural range. Understanding the patterns of genetic diversity in G. soja populations can help to inform conservation practices. Methods: In this study, we analyzed the genetic diversity and differentiation of G. soja at different sites and investigated the gene flow within the species. We obtained 147 G. soja accessions collected from 16 locations across the natural range of the species from China, Korea and Japan. Samples were analyzed using SLAF-seq (Specific-Locus Amplified Fragment Sequencing). Results: We obtained a total of 56,489 highly consistent SNPs. Our results suggested that G. soja harbors relatively high diversity and that populations of this species are highly differentiated. The populations harboring high genetic diversity, especially KR, should be considered first when devising conservation plans for the protection of G. soja, and in situ protection should be adopted in KR. G. soja populations from the Yangtze River, the Korean peninsula and northeastern China have a close relationship, although these areas are geographically disconnected. Other populations from north China clustered together. Analysis of gene flow suggested that historical migrations of G. soja may have occurred from the south northwards across the East-Asia land-bridge, but not across north China. All G. soja populations could be divided into one of two lineages, and these two lineages should be treated separately when formulating protection policies.

Mutant soluble ectodomain of fibroblast growth factor receptor-2 IIIc attenuates bleomycin-induced pulmonary fibrosis in mice.

We have developed a strong inhibitor (S252W mutant soluble ectodomain of fibroblast growth factor recptor-2 IIIc, msFGFR2) that binds FGFs strongly and blocks the activation of FGFRs. In vitro, msFGFR2 could inhibit the promoting effect of transforming growth factor (TGF)-ß1 on the proliferation of primary lung fibroblasts. In vivo, msFGFR2 alleviated lung fibrosis through inhibiting the expression of α-smooth muscle actin (SMA) and collagen deposit. In Western blotting of the right lung tissues and immunohistochemical assay, we found the level of p-FGFRs, p-mitogen activated protein kinase (MAPK) and p-Smad3 in the mice of bleomycin (BLM) group treated with msFGFR2 was down dramatically compared with the mice of BLM group, which suggested the activations of FGF and TGF-ß signals were blocked meanwhile. In summary, msFGFR2 attenuated BLM-induced fibrosis and is an attractive therapeutic candidate for human pulmonary fibrosis.

Genetic diversity and population structure: implications for conservation of wild soybean (Glycine soja Sieb. et Zucc) based on nuclear and chloroplast microsatellite variation.

Wild soybean (Glycine soja Sieb. et Zucc) is the most important germplasm resource for soybean breeding, and is currently subject to habitat loss, fragmentation and population decline. In order to develop successful conservation strategies, a total of 604 wild soybean accessions from 43 locations sampled across its range in China, Japan and Korea were analyzed using 20 nuclear (nSSRs) and five chloroplast microsatellite markers (cpSSRs) to reveal its genetic diversity and population structure. Relatively high nSSR diversity was found in wild soybean compared with other self-pollinated species, and the region of middle and lower reaches of Yangtze River (MDRY) was revealed to have the highest genetic diversity. However, cpSSRs suggested that Korea is a center of diversity. High genetic differentiation and low gene flow among populations were detected, which is consistent with the predominant self-pollination of wild soybean. Two main clusters were revealed by MCMC structure reconstruction and phylogenetic dendrogram, one formed by a group of populations from northwestern China (NWC) and north China (NC), and the other including northeastern China (NEC), Japan, Korea, MDRY, south China (SC) and southwestern China (SWC). Contrib analyses showed that southwestern China makes the greatest contribution to the total diversity and allelic richness, and is worthy of being given conservation priority.

The complete chloroplast genome of Lilium nepalense (Liliaceae).

Lilium nepalense is a useful plant species not only for its showy flowers but also has high medicinal value. In this study, the whole chloroplast genome of L. nepalense was sequenced for the first time. The genome size of L. nepalense, was 152,956bp, with typical tetragonal structure: one large single copy (82,573 bp), one small single copy (17,527 bp), and a pair of inverted repeat regions (IRs, 26,428 bp). The overall GC content was 37.0%. The complete genome contained 131 genes, including 85 protein-coding genes, 38 tRNA genes, and 8 rRNA genes. Phylogenetic analysis showed that L. nepalense was a close relationship between L. leucanthum and L. henryi. Phylogenetic analysis placed L. nepalense under the family Liliaceae.

The complete chloroplast genome and phylogenetic analysis of Gentiana arethusae Burkill (Gentianaceae) from China.

Gentiana arethusae Burkill is a perennial herb classified in the Gentianaceae. In this study, the complete chloroplast genome of G. arethusae was sequenced and analyzed. The chloroplast genome is 137,458 bp in length and encodes a total of 116 genes, including 71 protein-coding, 37 tRNA, and eight rRNA genes. The genome has a low GC content of 38.0%. Phylogenetic analysis of the genome of G. arethusae resolved it in a clade with Gentiana obconica and Gentiana veitchiorum. The complete chloroplast genome of G. arethusae will be helpful to study the genetic diversity and phylogenetics of the Gentianaceae.

The complete chloroplast genome of Lilium rosthornii Diels (Liliopsida: Liliaceae) from Hunan, China.

Lilium rosthornii is the perennial herbaceous bulbous plant belonging to the Lily of the Liliaceae, with high ornamental value and medicinal values. In this present study, we sequenced the complete chloroplast genome of Lilium rosthornii by Illumina Hiseq X Ten and PacBio RS technologies firstly. The genome size of L. rosthornii, was 152,242bp, with typical tetragonal structure: one large single-copy (LSC, 81,875 bp), one small single-copy (SSC, 17,553 bp), and a pair of inverted repeat regions (IRs, 26,407 bp). The overall GC content was 37.02%. The complete genome contained 131 genes, including 85 protein-coding genes, 38 tRNA genes, and eight rRNA genes. Phylogenetic analysis placed L. rosthornii under the family Liliaceae.

Analysis of Cucurbita ficifolia (Cucurbitaceae) chloroplast genome and its phylogenetic implications.

The figleaf gourd (Cucurbita ficifolia Bouché), is a member of the Cucurbitaceae. Figleaf gourd genotypes are exclusively used as a rootstock for cucumber owing to their high physiological compatibility with cucumber. In this study, the complete chloroplast (cp) genome of C. ficifolia was assembled. The cp genome of C. ficifolia was 157,631 bp in length, it consists of a pair of inverted repeats (IRa and IRb) regions (25,638 bp) separated by the large single-copy (LSC, 88,211 bp) and small single-copy (SSC, 18,144 bp) regions. The cp genome encodes 111 unique genes, including 80 protein-coding genes, 27 transfer RNA genes, and four ribosomal RNA genes. The overall GC content of C. ficifolia cp genome was 37.2%. The phylogenetic tree of Cucurbitaceae showed that C. ficifolia was clustered into genus Cucurbita and the bootstrap value is 100%.

The complete chloroplast genome of Lilium amoenum (Liliopsida: Liliaceae) from Yunnan, China.

Lilium amoenum E. H. Wilson ex Sealy is classified in Liliaceae, and it is an important ornamental plant with wonderful rose-red color and pleasant rose fragrance. In this study, we sequenced the complete chloroplast genome of L. amoenum by Illumina Hiseq X Ten and PacBio RS technologies. The genome size of L. amoenum is 152,280 bp, and displays a typical quadripartite structure: one large single-copy (LSC, 81,977 bp), one small single-copy (SSC, 17,539 bp), and a pair of inverted repeat regions (IRs, 26,382 bp). The overall GC content was 37.0%. The complete genome contained 131 genes, including 85 protein-coding genes, 38 tRNA genes, and 8 rRNA genes. Phylogenetic analysis showed that L. amoenum is closely related to L. taliense and L. bakerianum. The present study could afford crucial genetic information for further researches on the genus and related genera.

Characteristic and phylogenetic analyses of chloroplast genome for Mentha haplocalyx (Lamiaceae).

Mentha haplocalyx is an important medicinal and edible plant. Now, the complete chloroplast (cp) genome of M. haplocalyx was assembled and annotated. The cp genome of M. haplocalyx was 152,048 bp and contained two short inverted repeat regions (25,608 bp) which were separated by a small single copy region (17,654 bp) and a large single copyregion (83,179 bp). The cp genome encodes 113 unique genes, including 79 protein-coding genes, 30 transfer RNA genes and 4 ribosomal RNA genes. The topology of the phylogenetic tree showed that M. haplocalyx and M. spicata formed a monophyletic clade and clustered together with genus Dracocephalum.

Chloroplast genome for Crataegus pinnatifida (Rosaceae) and phylogenetic analyses with its coordinal species.

Crataegus pinnatifida is an important medicinal and edible plant. Now, the complete chloroplast (cp) genome of C. pinnatifida was assembled and annotated. The cp genome of C. pinnatifida was 159,898 bp and contained two short inverted repeat regions (26,540 bp) which were separated by a small single copy region (19,219 bp) and a large single copyregion (87,599 bp). The cp genome encodes 109 unique genes, including 75 protein-coding genes, 30 transfer RNA genes and 4 ribosomal RNA genes. The topology of the phylogenetic tree showed that C. pinnatifida has a close relationship with species Eriobotrya, Sorbus, Pyrus, Mulus, and Chaenomeles.

Phylogenetic analyses of Lycium chinense (Solanaceae) and its coordinal species applying chloroplast genome.

Lycium chinense is an important edible and medicinal plant. Now, the complete chloroplast (cp) genome of L. chinense was assembled based on the Illumina sequencing reads. The cp genome of L. chinense was 155,736 bp long and contained two short inverted repeat regions (25,469 bp), which were separated by a small single-copy region (18,206 bp) and a large single-copy region (86,592 bp). The cp genome encodes 113 unique genes, including 79 protein-coding genes, 30 transfer RNA genes, and 4 ribosomal RNA genes. The topology of the phylogenetic tree showed that L. chinense is closely clustered with species Lycium ruthenicum and Lycium barbarum.

Chloroplast genome and phylogenetic analyses of Morus alba (Moraceae).

Morus alba is an important medicinal plant that is used to treat human diseases. The complete chloroplast (cp) genome of M. alba was assembled based on the Illumina sequencing reads. The cp genome of M. alba was 159,290 bp and contained two short inverted repeat regions (25,690 bp) which were separated by a small single copy region (19,845 bp) and a large single copy region (88,065 bp). The cp genome encodes 111 unique genes, including 77 protein-coding genes, 30 transfer RNA genes and four ribosomal RNA genes. The topology of the phylogenetic tree showed that M. alba is closely clustered with species M. cathayana and M. mongolica.

Chloroplast genome and phylogenetic analyses of Poncirus trifoliata (Rutaceae).

Poncirus trifoliata is an important medicinal plant that is used to treat human diseases. In this study, the complete chloroplast (cp) genome of P. trifoliata was assembled based on the Illumina sequencing reads. The cp genome of P. trifoliata was 160,260 bp and contained two short inverted repeat regions (27,029 bp) which were separated by a small single copy region (18,760 bp) and a large single copy region (87,442 bp). The cp genome encodes 113 unique genes, including 79 protein-coding genes, 30 transfer RNA genes and 4 ribosomal RNA genes. The topology of the phylogenetic tree showed that P. trifoliata is closely clustered with genus Citrus.

Comparative analysis of four Zantedeschia chloroplast genomes: expansion and contraction of the IR region, phylogenetic analyses and SSR genetic diversity assessment.

The horticulturally important genus Zantedeschia (Araceae) comprises eight species of herbaceous perennials. We sequenced, assembled and analyzed the chloroplast (cp) genomes of four species of Zantedeschia (Z. aethiopica, Z. odorata, Z. elliottiana, and Z. rehmannii) to investigate the structure of the cp genome in the genus. According to our results, the cp genome of Zantedeschia ranges in size from 169,065 bp (Z. aethiopica) to 175,906 bp (Z. elliottiana). We identified a total of 112 unique genes, including 78 protein-coding genes, 30 transfer RNA (tRNA) genes and four ribosomal RNA (rRNA) genes. Comparison of our results with cp genomes from other species in the Araceae suggests that the relatively large sizes of the Zantedeschia cp genomes may result from inverted repeats (IR) region expansion. The sampled Zantedeschia species formed a monophylogenetic clade in our phylogenetic analysis. Furthermore, the long single copy (LSC) and short single copy (SSC) regions in Zantedeschia are more divergent than the IR regions in the same genus, and non-coding regions showed generally higher divergence than coding regions. We identified a total of 410 cpSSR sites from the four Zantedeschia species studied. Genetic diversity analyses based on four polymorphic SSR markers from 134 cultivars of Zantedeschia suggested that high genetic diversity (I = 0.934; Ne = 2.371) is present in the Zantedeschia cultivars. High genetic polymorphism from the cpSSR region suggests that cpSSR could be an effective tool for genetic diversity assessment and identification of Zantedeschia varieties.