Positive correlation between menthol content and in vitro menthol tolerance in Mentha arvensis L. cultivars

Menthol is a highly valued monoterpene produced by Japanese mint (Mentha arvensis) as a natural product with wide applications in cosmetics, confectionery, flavours, beverages and therapeutics. Selection of high menthol yielding genotypes is therefore the ultimate objective of all genetic improvement programmes in Mentha arvensis. A positive correlation was observed in the present study between menthol content in oils of evaluated genotypes and the level of tolerance to externally supplied menthol of explants of these genotypes in culture medium. The easy use of this relationship as a selectable biochemical marker opens the practical applicability of largescalein vitro screening of the germplasm, clones and breeders' material for selection of elite genotypes

Antibacterial efficacy of Jackfruit rag extract against clinically important pathogens and validation of its antimicrobial activity in Shigella dysenteriae infected Drosophila melanogaster infection model

513-522Exploration of alternative sources of antibacterial compounds is an important and possibly an effective solution to the current challenges in antimicrobial therapy. Plant derived wastes may offer one such alternative. Here, we investigated the antibacterial property of extract derived from a part of the Jackfruit (Artocarpus heterophyllus Lam.) called ‘rag’, generally considered as fruit waste. Morpho-physical characterization of the Jackfruit rag extract (JFRE) was performed using Gas-chromatography, where peaks indicative of furfural; pentanoic acid; and hexadecanoic acid were observed. In vitro biocompatibility of JFRE was performed using the MTT assay, which showed comparable cellular viability between extract-treated and untreated mouse fibroblast cells. Agar well disc diffusion assay exhibited JFRE induced zones of inhibition for a wide variety of laboratory and clinical strains of Gram-positive and Gram-negative bacteria. Analysis of electron microscope images of bacterial cells suggests that JFRE induces cell death by disintegration of the bacterial cell wall and precipitating intracytoplasmic clumping. The antibacterial activity of the JFREs was further validated in vivo using Shigella dysenteriae infected fly model, where JFRE pre-fed flies infected with S. dysenteriae had significantly reduced mortality compared to controls. JFRE demonstrates broad antibacterial property, both in vitro and in vivo, possibly by its activity on bacterial cell wall

Isolation of poly (A⁺)<i> </i>mRNA for downstream reactions from some medicinal and aromatic plants^†

197-201In the present protocol for extraction of RNA, hexadecyltrimethylammoniumbromide (CTAB) and insoluble polyvinylpyrrolidone were used followed by LiCl precipitation, CsCl ultracentrifugation and finally poly (A+) mRNA was isolated with the help of oligo(dT)-cellulose columns. The isolated poly (A+) mRNA was found to be suitable for cDNA-AFLP and suppression subtractive hybridization applications. It is a modified and consolidated protocol based on previously described methods for isolated steps and works better for medicinal and aromatic plants. High yield of poly (A+) mRNA coupled with its amenability for downstream reactions like RT-PCR. northern blotting and cDNA synthesis for library construction is a key feature of the present protocol

Rapid isolation of DNA from dry and fresh samples of plants producing large amounts of secondary metabolites and essential oils

The presence of certain metabolites has been observed to interfere with DNA isolation procedures and downstream reactions such as DNA restriction, amplification and cloning. The chemotypic heterogeneity among species may not permit optimal DNA yields with a single protocol, and thus, even closely related species may require different isolation protocols. Here we describe the essential steps of a rapid DNA isolation protocol that can be used for diverse medicinal and aromatic plants, which produce essential oils and secondary metabolites such as alkaloids, flavanoids, phenols, gummy polysaccharides, terpenes and quinones. The procedure is applicable to dry as well as fresh plant tissues. This protocol, in our experiments, permitted isolation of DNA from tissues of diverse plant species and produced fairly good yields. The isolated DNA proved amenable to PCR amplification and restriction digestion

Analysis of Homologous Regions of Small RNAs MIR397 and MIR408 Reveals the Conservation of Microsynteny among Rice Crop-Wild Relatives

MIRNAs are small non-coding RNAs that play important roles in a wide range of biological processes in plant growth and development. MIR397 (involved in drought, low temperature, and nitrogen and copper (Cu) starvation) and MIR408 (differentially expressed in response to environmental stresses such as copper, light, mechanical stress, dehydration, cold, reactive oxygen species, and drought) belong to conserved MIRNA families that either negatively or positively regulate their target genes. In the present study, we identified the homologs of MIR397 and MIR408 in Oryza sativa and its six wild progenitors, three non-Oryza species, and one dicot species. We analyzed the 100 kb segments harboring MIRNA homologs from 11 genomes to obtain a comprehensive view of their community evolution around these loci in the farthest (distant) relatives of rice. Our study showed that mature MIR397 and MIR408 were highly conserved among all Oryza species. Comparative genomics analyses also revealed that the microsynteny of the 100 kb region surrounding MIRNAs was only conserved in Oryza spp.; disrupted in Sorghum, maize, and wheat; and completely lost in Arabidopsis. There were deletions, rearrangements, and translocations within the 100 kb segments in Oryza spp., but the overall microsynteny of the region was maintained. The phylogenetic analyses of the precursor regions of all MIRNAs under study revealed a bimodal clade of common origin. This comparative analysis of miRNA involved in abiotic stress tolerance in plants provides a powerful tool for future Oryza research. Crop wild relatives (CWRs) offer multiple traits with potential to decrease the amount of yield loss owing to biotic and abiotic stresses. Using a comparative genomics approach, the exploration of CWRs as a source of tolerance to these stresses by understanding their evolution can be further used to leverage their yield potential

Menthol tolerant clones of Mentha arvensis: approach for in vitro selection of menthol rich genotypes

In vitro raised shoots of Mentha arvensis L. were screened for menthol tolerance level by growing them in media containing 0-100 μg ml<SUP>−1</SUP> menthol. A total of 2850 regenerated shoots were step wise screened for menthol tolerance at the concentrations of 50 μg ml<SUP>−1</SUP> followed by 60 and 70 μg ml<SUP>−1</SUP>. In this screening, only 30 individual regenerated shoots were able to survive. The clones from the primary screen were inoculated into rooting medium and, after rooting, transferred to pots in the greenhouse. Ultimately, these 30 menthol tolerant clones were multiplied and grown in the field in replicated plots of 2.5×2.5 m sizes. Twigs of 30 clones from the replicated trials were rechecked for tolerant phenotypes at a concentration of 70 μg ml<SUP>−1</SUP> menthol wherein, these survived even after 7 days (secondary screening). These clones were checked for oil and menthol content and were found to be better than the control plants. Out of these 30 plants, five tolerated 80 μg ml<SUP>−1</SUP> menthol (tertiary level screening) and were found to contain the highest amount of menthol per g leaf biomass. Molecular analysis through RAPD showed distinct variation in the profiles of these five plants, in comparison to the control. Using this method the relationship between the primer OPT 04, menthol tolerance and high menthol content character of the genotype was established. Further, a cultivar 'Saksham' was released from the selections by CIMAP for superior performance

Recessive Loci <i>Pps-1</i> and <i>OM</i> Differentially Regulate <i>PISTILLATA-1</i> and <i>APETALA3-1</i> Expression for Sepal and Petal Development in <i>Papaver somniferum</i>

<div><p>The involvement of PISTILLATA (PI) and APETALA (AP) transcription factors in the development of floral organs has previously been elucidated but little is known about their upstream regulation. In this investigation, two novel mutants generated in <i>Papaver somniferum</i> were analyzed - one with partially petaloid sepals and another having sepaloid petals. Progeny from reciprocal crosses of respective mutant parent genotypes showed a good fit to the monogenic Mendelian inheritance model, indicating that the mutant traits are likely controlled by the single, recessive nuclear genes named “<i>Pps-1</i>” and “<i>OM</i>” in the partially petaloid sepal and sepaloid petal phenotypes, respectively. Both paralogs of <i>PISTILLATA</i> (<i>PapsPI-1</i> and <i>PapsPI-3</i>) were obtained from the sepals and petals of <i>P. somniferum</i>. Ectopic expression of <i>PapsPI-1</i> in tobacco resulted in a partially petaloid sepal phenotype at a low frequency. Upregulation of <i>PapsPI-1</i> and <i>PapsAP3-1</i> in the petal and the petal part of partially petaloid sepal mutant and down-regulation of the same in sepaloid petal mutant indicates a differential pattern of regulation for flowering-related genes in various whorls. Similarly, it was found that the recessive mutation <i>OM</i> in sepaloid petal mutant downregulates <i>PapsPI-1</i> and <i>PapsAP3-1</i> transcripts. The recessive nature of the mutations was confirmed by the segregation ratios obtained in this analysis.</p></div

Comparison of quantitative expression levels of <i>PapsAP3-1</i> in the sepals of genotypes Pps-1 and I-14.

<p>The Y-axis represents relative quantities equilibrating the expression of <i>PapsAP3-1</i> in I-14 sepals as 1RQ value. Data represent mean <u>+</u> standard error of 3-5 biological replicates. On X-axis, S: sepal; Se: Petaloid-devoid part of the sepal and Pe: Petaloid part of the sepal.</p