Pharmacognostic Investigation of the Leaves of Mentha cordifolia and Its DNA Fingerprints

Objective: To perform microscopic investigation of Mentha cordifolia leaf and determine DNA fingerprint of the plant.Methods: Tissues of M. cordifolia, M. arvensis var piperascens and M. spicata were investigated under light microscope. DNA marker for M. cordifolia was established by amplification of the internal transcribed spacer (ITS) of nuclear ribosomal RNA gene using conserved plant sequences as primers and fragmentation with the restriction enzyme Bsm1.Results: The results showed the presence of uniseriate epidermal cells covered by a fine cuticle layer, glandular trichomes of multicellular type, capitate and peltate, and non-glandular trichomes. The fragmentation pattern of ITS of nuclear ribosomal RNA gene was applied as DNA marker for discrimination of M. cordifolia from M. arvensis var piperascens and M. spicata.Conclusion: Pharmacognostic investigation of M. cordifolia leaf exhibited the characteristic of Mentha species and the plant is distinguishable from M. arvensis var piperascens and M. spicata by DNA fingerprint pattern.Keywords: Mentha cordifolia, microscopic investigation, DNA marker, internal transcribed space

Integration of physical and genetic maps in apple confirms whole-genome and segmental duplications in the apple genome

A total of 355 simple sequence repeat (SSR) markers were developed, based on expressed sequence tag (EST) and bacterial artificial chromosome (BAC)-end sequence databases, and successfully used to construct an SSR-based genetic linkage map of the apple. The consensus linkage map spanned 1143 cM, with an average density of 2.5 cM per marker. Newly developed SSR markers along with 279 SSR markers previously published by the HiDRAS project were further used to integrate physical and genetic maps of the apple using a PCR-based BAC library screening approach. A total of 470 contigs were unambiguously anchored onto all 17 linkage groups of the apple genome, and 158 contigs contained two or more molecular markers. The genetically mapped contigs spanned ∼421 Mb in cumulative physical length, representing 60.0% of the genome. The sizes of anchored contigs ranged from 97 kb to 4.0 Mb, with an average of 995 kb. The average physical length of anchored contigs on each linkage group was ∼24.8 Mb, ranging from 17.0 Mb to 37.73 Mb. Using BAC DNA as templates, PCR screening of the BAC library amplified fragments of highly homologous sequences from homoeologous chromosomes. Upon integrating physical and genetic maps of the apple, the presence of not only homoeologous chromosome pairs, but also of multiple locus markers mapped to adjacent sites on the same chromosome was detected. These findings demonstrated the presence of both genome-wide and segmental duplications in the apple genome and provided further insights into the complex polyploid ancestral origin of the apple

Introduction of apple ANR genes into tobacco inhibits expression of both CHI and DFR genes in flowers, leading to loss of anthocyanin

Three genes encoding anthocyanidin reductase (ANR) in apple (Malus×domestica Borkh.), designated MdANR1, MdANR2a, and MdANR2b, have been cloned and characterized. MdANR1 shows 91% identity in coding DNA sequences with MdANR2a and MdANR2b, while MdANR2a and MdANR2b are allelic and share 99% nucleotide sequence identity in the coding region. MdANR1 and MdANR2 genes are located on linkage groups 10 and 5, respectively. Expression levels of both MdANR1 and MdANR2 genes are generally higher in yellow-skinned cv. Golden Delicious than in red-skinned cv. Red Delicious. Transcript accumulation of MdANR1 and MdANR2 genes in fruits gradually decreased throughout fruit development. Ectopic expression of apple MdANR genes in tobacco positively and negatively regulates the biosynthesis of proanthocyanidins (PAs) and anthocyanin, respectively, resulting in white, pale pink-coloured, and white/red variegated flowers. The accumulation of anthocyanin is significantly reduced in all tobacco transgenic flowers, while catechin and epicatechin contents in transgenic flowers are significantly higher than those in flowers of wild-type plants. The inhibition of anthocyanin synthesis in tobacco transgenic flowers overexpressing MdANR genes is probably attributed to down-regulation of CHALCONE ISOMERASE (CHI) and DIHYDROFLAVONOL-4-REDUCTASE (DFR) genes involved in the anthocyanin pathway. Interestingly, several transgenic lines show no detectable transcripts of the gene encoding leucoanthocyanidin reductase (LAR) in flowers, but accumulate higher levels of catechin in flowers of transgenic plants than those of wild-type plants. This finding suggests that the ANR gene may be capable of generating catechin via an alternative route, although this mechanism is yet to be further elucidated

Travel Tales of a Worldwide Weed: Genomic Signatures of Plantago major L. Reveal Distinct Genotypic Groups With Links to Colonial Trade Routes

Retracing pathways of historical species introductions is fundamental to understanding the factors involved in the successful colonization and spread, centuries after a species’ establishment in an introduced range. Numerous plants have been introduced to regions outside their native ranges both intentionally and accidentally by European voyagers and early colonists making transoceanic journeys; however, records are scarce to document this. We use genotyping-by-sequencing and genotype-likelihood methods on the selfing, global weed, Plantago major, collected from 50 populations worldwide to investigate how patterns of genomic diversity are distributed among populations of this global weed. Although genomic differentiation among populations is found to be low, we identify six unique genotype groups showing very little sign of admixture and low degree of outcrossing among them. We show that genotype groups are latitudinally restricted, and that more than one successful genotype colonized and spread into the introduced ranges. With the exception of New Zealand, only one genotype group is present in the Southern Hemisphere. Three of the most prevalent genotypes present in the native Eurasian range gave rise to introduced populations in the Americas, Africa, Australia, and New Zealand, which could lend support to the hypothesis that P. major was unknowlingly dispersed by early European colonists. Dispersal of multiple successful genotypes is a likely reason for success. Genomic signatures and phylogeographic methods can provide new perspectives on the drivers behind the historic introductions and the successful colonization of introduced species, contributing to our understanding of the role of genomic variation for successful establishment of introduced taxa.publishedVersio

Travel Tales of a Worldwide Weed: Genomic Signatures of Plantago major L. Reveal Distinct Genotypic Groups With Links to Colonial Trade Routes

Retracing pathways of historical species introductions is fundamental to understanding the factors involved in the successful colonization and spread, centuries after a species’ establishment in an introduced range. Numerous plants have been introduced to regions outside their native ranges both intentionally and accidentally by European voyagers and early colonists making transoceanic journeys; however, records are scarce to document this. We use genotyping-by-sequencing and genotype-likelihood methods on the selfing, global weed, Plantago major, collected from 50 populations worldwide to investigate how patterns of genomic diversity are distributed among populations of this global weed. Although genomic differentiation among populations is found to be low, we identify six unique genotype groups showing very little sign of admixture and low degree of outcrossing among them. We show that genotype groups are latitudinally restricted, and that more than one successful genotype colonized and spread into the introduced ranges. With the exception of New Zealand, only one genotype group is present in the Southern Hemisphere. Three of the most prevalent genotypes present in the native Eurasian range gave rise to introduced populations in the Americas, Africa, Australia, and New Zealand, which could lend support to the hypothesis that P. major was unknowlingly dispersed by early European colonists. Dispersal of multiple successful genotypes is a likely reason for success. Genomic signatures and phylogeographic methods can provide new perspectives on the drivers behind the historic introductions and the successful colonization of introduced species, contributing to our understanding of the role of genomic variation for successful establishment of introduced taxa.info:eu-repo/semantics/publishedVersio

The Orphan Gene ybjN Conveys Pleiotropic Effects on Multicellular Behavior and Survival of Escherichia coli

YbjN, encoding an enterobacteria-specific protein, is a multicopy suppressor of temperature sensitivity in the ts9 mutant strain of Escherichia coli. In this study, we further explored the role(s) of ybjN. First, we demonstrated that the ybjN transcript was about 10-fold lower in the ts9 strain compared to that of E. coli strain BW25113 (BW). Introduction of multiple copies of ybjN in the ts9 strain resulted in over-expression of ybjN by about 10-fold as compared to that of BW. These results suggested that temperature sensitivity of the ts9 mutant of E. coli may be related to expression levels of ybjN. Characterization of E. coli ybjN mutant revealed that ybjN mutation resulted in pleiotropic phenotypes, including increased motility, fimbriation (auto-aggregation), exopolysaccharide production, and biofilm formation. In contrast, over-expression of ybjN (in terms of multiple copies) resulted in reduced motility, fimbriation, exopolysaccharide production, biofilm formation and acid resistance. In addition, our results indicate that a ybjN-homolog gene from Erwinia amylovora, a plant enterobacterial pathogen, is functionally conserved with that of E. coli, suggesting similar evolution of the YbjN family proteins in enterobacteria. A microarray study revealed that the expression level of ybjN was inversely correlated with the expression of flagellar, fimbrial and acid resistance genes. Over-expression of ybjN significantly down-regulated genes involved in citric acid cycle, glycolysis, the glyoxylate shunt, oxidative phosphorylation, amino acid and nucleotide metabolism. Furthermore, over-expression of ybjN up-regulated toxin-antitoxin modules, the SOS response pathway, cold shock and starvation induced transporter genes. Collectively, these results suggest that YbjN may play important roles in regulating bacterial multicellular behavior, metabolism, and survival under stress conditions in E. coli. These results also suggest that ybjN over-expression-related temperature rescue of the ts9 mutant may be due to down-regulation of metabolic activity and activation of stress response genes in the ts9 mutant

Identifying and characterizing genes and transcription factors involved in coloration of apple fruit

Apple (Malus × domestica Borkh.) is one of the most highly cultivated fruit crops grown around the world and apple consumption has been increasing over the years. One of the most important determinants of fruit quality is skin color. Red coloration in apple fruit is attributed to anthocyanin accumulation. Anthocyanins are encoded by structural genes, in the anthocyanin biosynthetic pathway, which are highly regulated by transcription factors. In this thesis, studies were conducted to identify and characterize structural genes and associated transcription factors involved in the anthocyanin biosynthetic pathway. Three genes encoding apple anthocyanin reductase gene (MdANR) were isolated, designated as MdANR1, MdANR2a, and MdANR2b. It is found that MdANR2a, and MdANR2b are in fact allelic. MdANR1 and MdANR2 were mapped to the apple linkage map on linkage groups (LG) 10 and 5, respectively. The functionality of MdANR gene was investigated following its overexpression in tobacco and found to influence flower color pigmentation and pattern. Overexpression of MdANR influenced other genes in the flavonoid biosynthesis pathway by down-regulating chalcone isomerase (CHI), dihydroflavonol reductase (DFR), and leucoanthocyanidin reductase (LAR) genes. Moreover, the observed loss of flower color in transgenic tobacco lines was attributed to reduction of anthocyanin pigments. This was likely due to down-regulation of tobacco CHI and DFR genes that are important in anthocyanin production. In addition, a new floral pigmented pattern was generated by incomplete inhibition of anthocyanin production. As expected, the epicatechin accumulated at higher levels in transgenic tobacco than in wild-type tobacco. However, higher amounts of catechin but lower levels of LAR, responsible for synthesis of catechin, were found in transgenic lines when compared to wild-type tobacco. Thus, it has been proposed that ANR plays a redundant role to that of LAR. A novel MYB transcription factor (TF) gene, designated as MdMYB11, was isolated and genetically mapped onto LG15 of the apple genetic map. Alignment of deduced amino acid sequences of MdMYB11 to those of other R2R3 MYB TFs revealed that this new apple transcription factor contains the R2R3 conserved domain. Moreover, this TF is highly similar to Arabidopsis MYB subgroup 4, such as AtMYB3, 4, and 6, by which they negatively regulate genes involved in monolignol biosynthesis. Functional analysis of MdMYB11 was conducted via ectopic expression in tobacco. Expression of MdMYB11 increased anthocyanin production in tobacco flowers by inducing several anthocyanin biosynthesis pathway genes, particularly those of CHI, chalcone synthase (CHS), and UDP-glucose: flavonoid 3-o-glucosyltransferase (UFGT). In addition, this TF functioned as a repressor of both cinnamate-4-hydroxylase (C4H) and 4-coumaroyl:CoA-ligase (4CL) genes, both important in lignin biosynthesis, and possibly contributing to modulation of floral morphogenesis. Moreover, transgenic flowers had longer styles than those of wild-type flowers, suggesting that the MdMYB11 gene might be involved in pistil development. New candidate TF genes regulating apple fruit coloration were identified following global gene expression analysis of the apple transcriptome using an apple microarray. Comparison of gene expression in fruit peel of apple cv. Red Delicious subjected to continuous ‘dark treatment’ versus dark-grown fruit subjected to ‘14 h-light-exposure’ identified 815 genes that were modulated. Following annotation (to the Arabidopsis Gene Ontology), these genes were classified into 19 categories, and were mostly involved in primary metabolism (17%) and transcription (12%). Of these, 18 genes encoded for putative TFs. Further identification of color-related TFs was conducted by comparison of expression profiles of fruit of red skinned apple cv. Red Delicious and non-red skinned apple cv. Golden Delicious, and using quantitative real-time (RT)-PCR (qRT-PCR). Two putative TF genes were found to be expressed at higher levels in fruit of ‘Red Delicious’ than that in ‘Golden Delicious’, thus suggesting that these TFs might be involved in fruit coloration. Altogether, these findings have provided novel information and knowledge of the role(s) of genes and transcription factors involved in the anthocyanin biosynthesis pathway. Moreover, the regulator mechanism of fruit coloration has been further elucidated following transcriptome analysis of the apple genome and functional analysis of selected genes and transcription factors

Chemical compositions, antioxidant, antimicrobial, and mosquito larvicidal activity of Ocimum americanum L. and Ocimum basilicum L. leaf essential oils

Abstract Background Ocimum americanum L. (O. americanum) and Ocimum basilicum L. (O. basilicum) are highly valued aromatic medicinal plants. Their leaves are widely used as spices in traditional cuisine. Their essential oils (EOs) are extensively used in food, cosmetic, and pharmaceutical industries. This study aimed to investigate the main chemical profiles of O. americanum and O. basilicum leaf EOs and assess their effects on antibacterial, antioxidant, and larvicidal properties. Methods EOs were extracted from the leaves of O. basilicum and O. americanum using steam distillation in a Clevenger-type apparatus. The chemical constituents of the EOs were analyzed using gas chromatography–mass spectrometry. 2,2-Diphenyl-1-picrylhydrazyl (DPPH), ferric reducing antioxidant power (FRAP), and metal-chelating techniques were used to assess the free-radical scavenging capability of the oils. The extracted oils were also tested for their antibacterial activities via a disk-diffusion test and the broth microdilution method. Furthermore, the mosquito larvicidal (Aedes aegypti) activity was tested using standard protocols. Results Camphor (33.869%), limonene (7.215%), longifolene (6.727%), caryophyllene (5.500%), and isoledene (5.472%) were the major compounds in O. americanum leaf EO. The EO yield was 0.4%, and citral (19.557%), estragole (18.582%) camphor (9.224%) and caryophyllene (3.009%) were the major compounds found among the 37 chemical constituents identified in O. basilicum oil. O. basilicum exhibited a more potent antioxidant activity in DPPH, FRAP, and 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid tests than O. americanum. The zones of inhibition and minimum inhibitory concentration of the oils in the microdilution and disk diffusion methods were 8.00 ± 0.19 mm to 26.43 ± 2.19 mm and 3.12–100 µg/mL, respectively. At 400 ppm, O. basilicum and O. americanum EOs demonstrated larvicidal activity, with mortality ratios of 73.60% ± 0.89% and 78.00% ± 1.00%, respectively. Furthermore, after 30 min of exposure to O. americanum and O. basilicum EOs, the larval death rates were 73.60% ± 0.89% and 78.00% ± 1.00%, respectively. Conclusions The findings revealed that the EOs extracted from the leaves of O. basilicum and O. americanum exhibited reasonable antioxidant, antibacterial, and mosquito larvicidal potentials, and can be used as alternative medicine for the treatment of human health and larvicidal mosquito control

Enhanced Physicochemical Stability of the L-DOPA Extract of <i>Mucuna pruriens</i> Seeds by Adding <i>Phyllanthus emblica</i>

Levodopa (L-DOPA) is an essential drug for the treatment of Parkinson’s disease. Currently, L-DOPA can be produced by chemical synthesis and can also be found naturally in many herbs, especially Mucuna Pruriens (MP). According to clinical research, the MP extract containing L-DOPA for the treatment of Parkinson’s disease could reduce side effects more than the synthetic one. Unfortunately, MP extracts can be easily degraded. Changes in physical and chemical properties such as the appearance (color, melt, solid lump) and the reduction of L-DOPA content in the extract were commonly observed. Therefore, it is necessary to develop an extraction procedure to stabilize the extract of L-DOPA. This study attempted to enhance the extraction process by modifying the traditional acidification approach using hydrochloric acid, citric acid, or ascorbic acid. According to the stability test results, using Phyllanthus emblica water (PEW) as a solvent improved the preservative properties more than other solvents. The color of the PEW-MP powder changed slightly after 12 months of accelerated storage, but the amount of L-DOPA remained the highest (73.55%). Moreover, L-DOPA was only detected in MP and PEW-MP, but not PEW alone (the HPTLC chromatogram at Rf 0.48 and the HPLC chromatogram at Rt 6.0 min). The chemical profiles of PEW and L-DOPA observed in the chromatograms indicated that they are independently separated. As a result, they can be applied to a quality control process. Therefore, PEW was proven to be a powerful solvent for L-DOPA herbal extract that could be readily used as a raw material for herbal products