Molecular mechanism of miRNA mediated biosynthesis of secondary metabolites in medicinal plants.

Plant secondary metabolites are important raw materials for the pharmaceutical industry, and their biosynthetic processes are subject to diverse and precise regulation by miRNA. The identification of miRNA molecules in medicinal plants and exploration of their mechanisms not only contribute to a deeper understanding of the molecular genetic mechanisms of plant growth, development and resistance to stress, but also provide a theoretical basis for elucidating the pharmacological effects of authentic medicinal materials and constructing bioreactors for the synthesis of medicinal secondary metabolite components. This paper summarizes the research reports on the discovery of miRNA in medicinal plants and their regulatory mechanisms on the synthesis of secondary metabolites by searching the relevant literature in public databases. It summarizes the currently discovered miRNA and their functions in medicinal plants, and summarizes the molecular mechanisms regulating the synthesis and degradation of secondary metabolites. Furthermore, it provides a prospect for the research and development of medicinal plant miRNA. The compiled information contributes to a comprehensive understanding of the research progress on miRNA in medicinal plants and provides a reference for the industrial development of related secondary metabolite biosynthesis.

A century of studying plant secondary metabolism-From "what?" to "where, how, and why?"

Over the past century, early advances in understanding the identity of the chemicals that collectively form a living plant have led scientists to deeper investigations exploring where these molecules localize, how they are made, and why they are synthesized in the first place. Many small molecules are specific to the plant kingdom and have been termed plant secondary metabolites, despite the fact that they can play primary and essential roles in plant structure, development, and response to the environment. The past 100 yr have witnessed elucidation of the structure, function, localization, and biosynthesis of selected plant secondary metabolites. Nevertheless, many mysteries remain about the vast diversity of chemicals produced by plants and their roles in plant biology. From early work characterizing unpurified plant extracts, to modern integration of 'omics technology to discover genes in metabolite biosynthesis and perception, research in plant (bio)chemistry has produced knowledge with substantial benefits for society, including human medicine and agricultural biotechnology. Here, we review the history of this work and offer suggestions for future areas of exploration. We also highlight some of the recently developed technologies that are leading to ongoing research advances.

Selenium nanoparticles conferred drought tolerance in tomato plants by altering the transcription pattern of microRNA-172 (miR-172), bZIP, and CRTISO genes, upregulating the antioxidant system, and stimulating secondary metabolism.

Drought stress is one of the major limiting factors for the production of tomato in Iran. In this study, the efficiency of selenate and Se nanoparticle (SeNP) foliar application on tomato plants was assessed to vestigate mitigating the risk associated with water-deficit conditions. Tomato plants were treated with SeNPs at the concentrations of 0 and 4 mg L-1; after the third sprays, the plants were exposed to water-deficit conditions. The foliar spraying with SeNPs not only improved growth, yield, and developmental switch to the flowering phase but also noticeably mitigated the detrimental risk associated with the water-deficit conditions. Gene expression experiments showed a slight increase in expression of microRNA-172 (miR-172) in the SeNP-treated plants in normal irrigation, whereas miR-172 displayed a downregulation trend in response to drought stress. The bZIP transcription factor and CRTISO genes were upregulated following the SeNP and drought treatments. Drought stress significantly increased the H2O2 accumulation that is mitigated with SeNPs. The foliar spraying with Se or SeNPs shared a similar trend to alleviate the negative effect of drought stress on the membrane integrity. The applied supplements also conferred drought tolerance through noticeable improvements in the non-enzymatic (ascorbate and glutathione) and enzymatic (catalase and peroxidase) antioxidants. The SeNP-mediated improvement in drought stress tolerance correlated significantly with increases in the activity of phenylalanine ammonia-lyase, proline, non-protein thiols, and flavonoid concentrations. SeNPs also improved the fruit quality regarding K, Mg, Fe, and Se concentrations. It was concluded that foliar spraying with SeNPs could mitigate the detrimental risk associated with the water-deficit conditions.

Role of bZIP transcription factors in the regulation of plant secondary metabolism.

MAIN CONCLUSION: This study provides an overview of the structure, classification, regulatory mechanisms, and biological functions of the basic (region) leucine zipper transcription factors and their molecular mechanisms in flavonoid, terpenoid, alkaloid, phenolic acid, and lignin biosynthesis. Basic (region) leucine zippers (bZIPs) are evolutionarily conserved transcription factors (TFs) in eukaryotic organisms. The bZIP TFs are widely distributed in plants and play important roles in plant growth and development, photomorphogenesis, signal transduction, resistance to pathogenic microbes, biotic and abiotic stress, and secondary metabolism. Moreover, the expression of bZIP TFs not only promotes or inhibits the accumulation of secondary metabolites in medicinal plants, but also affects the stress response of plants to the external adverse environment. This paper describes the structure, classification, biological function, and regulatory mechanisms of bZIP TFs. In addition, the molecular mechanism of bZIP TFs regulating the biosynthesis of flavonoids, terpenoids, alkaloids, phenolic acids, and lignin are also elaborated. This review provides a summary for in-depth study of the molecular mechanism of bZIP TFs regulating the synthesis pathway of secondary metabolites and plant molecular breeding, which is of significance for the generation of beneficial secondary metabolites and the improvement of plant varieties.

Lavandula Species, Their Bioactive Phytochemicals, and Their Biosynthetic Regulation.

Lavandula species are one of the most useful aromatic and medicinal plants and have great economic potential. The phytopharmaceutical contribution of the secondary metabolites of the species is unquestionable. Most recent studies have been focusing on the elucidation of the genetic background of secondary metabolite production in lavender species. Therefore, knowledge of not only genetic but especially epigenetic mechanisms for the regulation of secondary metabolites is necessary for the modification of those biosynthesis processes and the understanding of genotypic differences in the content and compositional variability of these products. The review discusses the genetic diversity of Lavandula species in relation to the geographic area, occurrence, and morphogenetic factors. The role of microRNAs in secondary-metabolites biosynthesis is described.

Metabolic and Transcriptional Stress Memory in Sorbus pohuashanensis Suspension Cells Induced by Yeast Extract.

Plant stress memory can provide the benefits of enhanced protection against additional stress exposure. Here, we aimed to explore the responses of recurrent and non-recurrent yeast extract (YE) stresses in Sorbus pohuashanensis suspension cells (SPSCs) at metabolomics and transcriptional levels. Biochemical analyses showed that the cell wall integrity and antioxidation capacity of SPSCs in the pretreated group were evidently improved. Metabolic analysis showed that there were 39 significantly altered metabolites in the pretreated group compared to the non-pretreated group. Based on the transcriptome analysis, 219 differentially expressed genes were obtained, which were highly enriched in plant-pathogen interaction, circadian rhythm-plant, oxidative phosphorylation, and phenylpropanoid biosynthesis. Furthermore, the correlation analysis of the transcriptome and metabolome data revealed that phenylpropanoid biosynthesis involved in the production of biphenyl phytoalexins may play a critical role in the memory response of SPSC to YE, and the key memory genes were also identified, including PAL1, BIS1, and BIS3. Collectively, the above results demonstrated that the memory responses of SPSC to YE were significant in almost all levels, which would be helpful for better understanding the adaptation mechanisms of medicinal plants in response to biotic stress, and laid a biotechnological foundation to accumulate favorable antimicrobial drug candidates from plant suspension cells.

The Current Developments in Medicinal Plant Genomics Enabled the Diversification of Secondary Metabolites' Biosynthesis.

Medicinal plants produce important substrates for their adaptation and defenses against environmental factors and, at the same time, are used for traditional medicine and industrial additives. Plants have relatively little in the way of secondary metabolites via biosynthesis. Recently, the whole-genome sequencing of medicinal plants and the identification of secondary metabolite production were revolutionized by the rapid development and cheap cost of sequencing technology. Advances in functional genomics, such as transcriptomics, proteomics, and metabolomics, pave the way for discoveries in secondary metabolites and related key genes. The multi-omics approaches can offer tremendous insight into the variety, distribution, and development of biosynthetic gene clusters (BGCs). Although many reviews have reported on the plant and medicinal plant genome, chemistry, and pharmacology, there is no review giving a comprehensive report about the medicinal plant genome and multi-omics approaches to study the biosynthesis pathway of secondary metabolites. Here, we introduce the medicinal plant genome and the application of multi-omics tools for identifying genes related to the biosynthesis pathway of secondary metabolites. Moreover, we explore comparative genomics and polyploidy for gene family analysis in medicinal plants. This study promotes medicinal plant genomics, which contributes to the biosynthesis and screening of plant substrates and plant-based drugs and prompts the research efficiency of traditional medicine.

Diverse roles of MYB transcription factors in regulating secondary metabolite biosynthesis, shoot development, and stress responses in tea plants (Camellia sinensis).

Tea (Camellia sinensis) is concocted from tea plant shoot tips that produce catechins, caffeine, theanine, and terpenoids, which collectively determine the rich flavors and health benefits of the infusion. However, little is known about the integrated regulation of shoot tip development and characteristic secondary metabolite biosynthesis in tea plants. Here, we demonstrate that MYB transcription factors (TFs) play key and yet diverse roles in regulating leaf and stem development, secondary metabolite biosynthesis, and environmental stress responses in tea plants. By integrating transcriptomic and metabolic profiling data in different tissues at a series of developmental stages or under various stress conditions, alongside biochemical and genetic analyses, we predicted the MYB TFs involved in regulating shoot development (CsMYB2, 98, 107, and 221), epidermal cell initiation (CsMYB184, 41, 139, and 219), stomatal initiation (CsMYB113 and 153), and the biosynthesis of flavonoids (including catechins, anthocyanins, and flavonols; CsMYB8 and 99), caffeine (CsMYB85 and 86), theanine (CsMYB9 and 49), carotenoids (CsMYB110), mono-/sesquiterpenoid volatiles (CsMYB68, 147, 148, and 193), lignin (CsMYB164 and 192), and indolic compounds (CsMYB139, 162, and 198), as well as the MYB TFs that are likely involved in hormone signaling-mediated environmental stress and defense responses. We characterized the functions of some key MYBs in regulating flavonoid and carotenoid biosynthesis for tea quality and flavor. This study provides a cross-family analysis of MYBs in tea alongside new insights into the coordinated regulation of tea plant shoot development and secondary metabolism, paving the way towards understanding of tea quality trait formation and genetic improvement of quality tea plants.

Sexual Crossing, Chromosome-Level Genome Sequences, and Comparative Genomic Analyses for the Medicinal Mushroom Taiwanofungus Camphoratus (Syn. Antrodia Cinnamomea, Antrodia Camphorata).

Taiwanofungus camphoratus mushrooms are a complementary and alternative medicine for hangovers, cancer, hypertension, obesity, diabetes, and inflammation. Though Taiwanofungus camphoratus has attracted considerable biotechnological and pharmacological attention, neither classical genetic nor genomic approaches have been properly established for it. We isolated four sexually competent monokaryons from two T. camphoratus dikaryons used for the commercial cultivation of orange-red (HC1) and milky-white (SN1) mushrooms, respectively. We also sequenced, annotated, and comparatively analyzed high-quality and chromosome-level genome sequences of these four monokaryons. These genomic resources represent a valuable basis for understanding the biology, evolution, and secondary metabolite biosynthesis of this economically important mushrooms. We demonstrate that T. camphoratus has a tetrapolar mating system and that HC1 and SN1 represent two intraspecies isolates displaying karyotypic variation. Compared with several edible mushroom model organisms, T. camphoratus underwent a significant contraction in the gene family and individual gene numbers, most notably for plant, fungal, and bacterial cell-wall-degrading enzymes, explaining why T. camphoratus mushrooms are rare in natural environments, are difficult and time-consuming to artificially cultivate, and are susceptible to fungal and bacterial infections. Our results lay the foundation for an in-depth T. camphoratus study, including precise genetic manipulation, improvements to mushroom fruiting, and synthetic biology applications for producing natural medicinal products. IMPORTANCETaiwanofungus camphoratus (Tc) is a basidiomycete fungus that causes brown heart rot of the aromatic tree Cinnamomum kanehirae. The Tc fruiting bodies have been used to treat hangovers, abdominal pain, diarrhea, hypertension, and other diseases first by aboriginal Taiwanese and later by people in many countries. To establish classical genetic and genomic approaches for this economically important medicinal mushroom, we first isolated and characterized four sexually competent monokaryons from two dikaryons wildly used for commercial production of Tc mushrooms. We applied PacBio single molecule, real-time sequencing technology to determine the near-completed genome sequences of four monokaryons. These telomere-to-telomere and gapless haploid genome sequences reveal all genomic variants needed to be studied and discovered, including centromeres, telomeres, retrotransposons, mating type loci, biosynthetic, and metabolic gene clusters. Substantial interspecies diversities are also discovered between Tc and several other mushroom model organisms, including Agrocybe aegerita, Coprinopsis cinerea, and Schizophyllum commune, and Ganoderma lucidum.

Targeting Mitochondria by Plant Secondary Metabolites: A Promising Strategy in Combating Parkinson's Disease.

Parkinson's disease (PD) is one of the most prevalent and debilitating neurodegenerative conditions, and is currently on the rise. Several dysregulated pathways are behind the pathogenesis of PD; however, the critical targets remain unclear. Accordingly, there is an urgent need to reveal the key dysregulated pathways in PD. Prevailing reports have highlighted the importance of mitochondrial and cross-talked mediators in neurological disorders, genetic changes, and related complications of PD. Multiple pathophysiological mechanisms of PD, as well as the low efficacy and side effects of conventional neuroprotective therapies, drive the need for finding novel alternative agents. Recently, much attention has been paid to using plant secondary metabolites (e.g., flavonoids/phenolic compounds, alkaloids, and terpenoids) in the modulation of PD-associated manifestations by targeting mitochondria. In this line, plant secondary metabolites have shown promising potential for the simultaneous modulation of mitochondrial apoptosis and reactive oxygen species. This review aimed to address mitochondria and multiple dysregulated pathways in PD by plant-derived secondary metabolites.

Transcriptome analysis reveals the molecular mechanisms of response to an emergent yellow-flower disease in green Chinese prickly ash (Zanthoxylum schinifolium).

Chinese prickly ash (Zanthoxylum) is extensively used as spice and traditional medicine in eastern Asian countries. Recently, an emergent yellow-flower disease (YFD) break out in green Chinese prickly ash (Zanthoxylum schinifolium, Qinghuajiao in Chinese) at Chongqing municipality, and then leads to a sharp reduction in the yield of Qinghuajiao, and thus results in great economic losses for farmers. To address the molecular response for the emergent YFD of Qinghuajiao, we analyzed the transcriptome of 12 samples including the leaves and inflorescences of asymptomatic and symptomatic plants from three different towns at Chongqing by high-throughput RNA-Seq technique. A total of 126,550 genes and 229,643 transcripts were obtained, and 21,054 unigenes were expressed in all 12 samples. There were 56 and 164 different expressed genes (DEGs) for the AL_vs_SL (asymptomatic leaf vs symptomatic leaf) and AF_vs_SF (asymptomatic flower vs symptomatic flower) groups, respectively. The results of KEGG analysis showed that the "phenylpropanoid biosynthesis" pathway that related to plant-pathogen interaction were found in AL_vs_SL and AF_vs_SF groups, and the "Plant-pathogen interaction" found in AF_vs_SF group, implying that this Qinghuajiao YFD might cause by plant pathogen. Interestingly, we detected 33 common unigenes for the 2 groups, and almost these unigenes were up-regulated in the symptomatic plants. Moreover, most of which were homologs to virus RNA, the components of viruses, implying that this YFD was related to virus. Our results provided a primary molecular basis for the prevention and treatment of YFD of Qinghuajiao trees.

Comparative transcriptome analysis of tubers, stems, and flowers of Gastrodia elata Blume reveals potential genes involved in the biosynthesis of phenolics.

Orchidaceae, well known for its fascinating flowers, is one of the largest and most diverse families of flowering plants. There are many kinds of plants in this family; these are distributed practically globally and have high ornamental and medicinal values. Gastrodia elata Blume, a traditional Chinese medicinal herb, is a rootless and leafless achlorophyllous orchid. Phenolic compounds are considered to be the major bioactive constituents in G. elata, with antioxidant, antiangiogenic, neuroprotective, antidepressant, anxiolytic, and sedative activities. In this study, we determined the contents of six main phenolic components in tubers, stems and flowers from G. elata. Meanwhile, the transcriptomes of the tuber, stem and flower tissues of G. elata were obtained using the BGISEQ-500 platform. A total of 58.29 Gb of data and 113,067 unigenes were obtained, of which 74,820 unigenes were functionally annotated against seven public databases. Differentially expressed genes between tuber, stem and flower tissues were identified. A total of 76 DEGs encoding eight key enzymes were identified as candidate genes involved in the biosynthesis of phenolics in G. elata. For further validation, the expression levels of unigenes were measured using quantitative real-time PCR. Our results greatly enrich the transcriptomic data of G. elata and provide valuable information for the identification of candidate genes involved in the biosynthesis of secondary metabolites.

Ecophysiological traits differentially modulate secondary metabolite accumulation and antioxidant properties of tea plant [Camellia sinensis (L.) O. Kuntze].

Owing to the diverse growing habitats, ecophysiology might have a regulatory impact on characteristic chemical components of tea plant. This study aimed to explore natural variations in the ecophysiological traits within seasons and the corresponding multifaceted biochemical responses given by the gene pool of 22 tea cultivars. Leaf temperature and intercellular carbon concentration (Ci), which varies as a function of transpiration and net photosynthesis respectively, have significant impact on the biochemical traits of the leaf. Occurrence of H2O2, in leaves, was associated to Ci that in turn influenced the lipid peroxidation. With the increment of Ci, total phenolics, epicatechin gallate (ECG), reducing power, and radical scavenging activity is lowered but total catechin and non-gallylated catechin derivatives (e.g. epicatechin or EC, epigallocatechin or EGC) are elevated. Leaf temperature is concomitantly associated (p ≤ 0.01) with phenolics, flavonoids, proanthocyanidin, tannin content, reducing power, iron chelation and free radical scavenging activities. Increased phenolic concentration in leaf cells, conceivably inhibit photosynthesis and moreover, gallic acid, thereafter conjugated to catechin derivatives. This study shed light on the fundamental information regarding ecophysiological impact on the quality determining biochemical characteristics of tea, which on further validation, might ascertain the genotype selection paradigm toward climate smart cultivation.

Genome sequencing and annotation and phylogenomic analysis of the medicinal mushroom Amauroderma rugosum, a traditional medicinal species in the family Ganodermataceae.

Amauroderma rugosum is one of the traditional Chinese medicinal mushrooms and is used to reduce inflammation, treat diuretic and upset stomach, and prevent cancer. Here, we present a genomic resource of Amauroderma rugosum (ACCC 51706) for further understanding its biology and exploration of the synthesis pathway of bioactive compounds. Genomic DNA was extracted and then subjected to Illumina HiSeq X Ten and PacBio Sequel I sequencing. The final genome is 40.66 Mb in size, with an N50 scaffold size of 36.6 Mb, and encodes 10 181 putative predicted genes. Among them, 6931 genes were functionally annotated. Phylogenomic analysis suggested that A. rugosum and Ganoderma sinense were not clustered together into a group and the latter was grouped with the Polyporaceae. Further, we also identified 377 carbohydrate-active enzymes (CAZymes) and 15 secondary metabolite biosynthetic gene clusters. This is the first genome-scale assembly and annotation for an Amauroderma species. The identification of novel secondary metabolite biosynthetic gene clusters would promote pharmacological research and development of novel bioactive compounds in the future.

The biosynthetic pathway of potato solanidanes diverged from that of spirosolanes due to evolution of a dioxygenase.

Potato (Solanum tuberosum), a worldwide major food crop, produces the toxic, bitter tasting solanidane glycoalkaloids α-solanine and α-chaconine. Controlling levels of glycoalkaloids is an important focus on potato breeding. Tomato (Solanum lycopersicum) contains a bitter spirosolane glycoalkaloid, α-tomatine. These glycoalkaloids are biosynthesized from cholesterol via a partly common pathway, although the mechanisms giving rise to the structural differences between solanidane and spirosolane remained elusive. Here we identify a 2-oxoglutarate dependent dioxygenase, designated as DPS (Dioxygenase for Potato Solanidane synthesis), that is a key enzyme for solanidane glycoalkaloid biosynthesis in potato. DPS catalyzes the ring-rearrangement from spirosolane to solanidane via C-16 hydroxylation. Evolutionary divergence of spirosolane-metabolizing dioxygenases contributes to the emergence of toxic solanidane glycoalkaloids in potato and the chemical diversity in Solanaceae.

Characterizing microRNAs and their targets in different organs of Camellia sinensis var. assamica.

To comprehensively annotate miRNAs and their targets in tea plant, Camellia sinensis, we sequenced small and messenger RNAs of 9 samples of Camellia sinensis var. assamica (YK-10), a diploid elite cultivar widely grown in southwest China. In order to identify targets of miRNAs, we sequenced two degradome sequencing profiles from leaves and roots of YK-10, respectively. By analyzing the small RNA-Seq profiles, we newly identified 137 conserved miRNAs and 23 species specific miRNAs in the genome of YK-10, which significantly improved the annotation of miRNAs in tea plant. Approximately 2000 differently expressed genes were identified when comparing RNA-Seq profiles of any two of the three organs selected in the study. Totally, more than 5000 targets of conserved miRNAs were identified in the two degradome profiles. Furthermore, our results suggest that a few miRNAs play roles in the biosynthesis pathways of theanine, caffeine and flavonoid. These results enhance our understanding of small RNA guided gene regulations in different organs of tea plant.

A predicted protein functional network aids in novel gene mining for characteristic secondary metabolites in tea plant (Camellia sinensis).

Modeling a protein functional network in concerned species is an efficient approach for identifying novel genes in certain biological pathways. Tea plant (Camellia sinensis) is an important commercial crop abundant in numerous characteristic secondary metabolites (e.g., polyphenols, alkaloids, alkaloids) that confer tea quality and health benefits. Decoding novel genes responsible for tea characteristic components is an important basis for applied genetic improvement and metabolic engineering. Herein, a high-quality protein functional network for tea plant (TeaPoN) was predicted using cross-species protein functional associations transferring and integration combined with a stringent biological network criterion control. TeaPoN contained 31,273 nonredundant functional interactions among 6,634 tea proteins (or genes), with general network topological properties such as scale-free and small-world. We revealed the modular organization of genes related to the major three tea characteristic components (theanine, caffeine, catechin) in TeaPoN, which served as strong evidence for the utility of TeaPoN in novel gene mining. Importantly, several case studies regarding gene identification for tea characteristic components were presented. To aid in the use of TeaPoN, a concise web interface for data deposit and novel gene screening was developed (http://teapon.wchoda.com). We believe that TeaPoN will serve as a useful platform for functional genomics studies associated with characteristic secondary metabolites in tea plant.

In silico and in vitro neutralization of PLA2 activity of Daboxin P by butein, mimosine and bakuchiol.

Medicinal plants have always been used for snakebite treatment by traditional healers but they lack scientific evidence of action. However secondary metabolites of such plants have been explored and found to inhibit the toxic effect of venom proteins. Literature survey from 2003 to 2019 resulted in identification of 251 secondary metabolites with such properties. In silico docking studies of these metabolites with modelled structure of Daboxin P, a PLA2 from Indian Daboia russelii revealed that butein, mimosine and bakuchiol bind to Daboxin P with high affinity. Butein interacted with the catalytic triad but mimosine and bakuchiol interacted with the Ca2+ binding residues of Daboxin P. In vitro validation showed that the molecules inhibited the sPLA2 activity of Daboxin P. Interestingly, mimosine and bakuchiol could also neutralize the anti-coagulatory activity of Daboxin P. Further, it was observed that butein and mimosine could neutralize the PLA2 activity of Indian big four venoms dose dependently. On the other hand, mimosine and bakuchiol could also neutralize the pro/anti-coagulatory effect of big four crude venom. Thus, in this study, three molecules have been identified which can neutralize the PLA2 activity and pro/anti-coagulatory effect of Daboxin P as well as crude venom of big four.

De novo transcriptome analysis of the critically endangered alpine Himalayan herb Nardostachys jatamansi reveals the biosynthesis pathway genes of tissue-specific secondary metabolites.

The study is the first report on de novo transcriptome analysis of Nardostachys jatamansi, a critically endangered medicinal plant of alpine Himalayas. Illumina GAIIx sequencing of plants collected during end of vegetative growth (August) yielded 48,411 unigenes. 74.45% of these were annotated using UNIPROT. GO enrichment analysis, KEGG pathways and PPI network indicated simultaneous utilization of leaf photosynthates for flowering, rhizome fortification, stress response and tissue-specific secondary metabolites biosynthesis. Among the secondary metabolite biosynthesis genes, terpenoids were predominant. UPLC-PDA analysis of in vitro plants revealed temperature-dependent, tissue-specific differential distribution of various phenolics. Thus, as compared to 25 °C, the phenolic contents of both leaves (gallic acid and rutin) and roots (p-coumaric acid and cinnamic acid) were higher at 15 °C. These phenolics accounted for the therapeutic properties reported in the plant. In qRT-PCR of in vitro plants, secondary metabolite biosynthesis pathway genes showed higher expression at 15 °C and 14 h/10 h photoperiod (conditions representing end of vegetative growth period). This provided cues for in vitro modulation of identified secondary metabolites. Such modulation of secondary metabolites in in vitro systems can eliminate the need for uprooting N. jatamansi from wild. Hence, the study is a step towards effective conservation of the plant.

A New High-Quality Draft Genome Assembly of the Chinese Cordyceps Ophiocordyceps sinensis.

Ophiocordyceps sinensis (Berk.) is an entomopathogenic fungus endemic to the Qinghai-Tibet Plateau. It parasitizes and mummifies the underground ghost moth larvae, then produces a fruiting body. The fungus-insect complex, called Chinese cordyceps or "DongChongXiaCao," is not only a valuable traditional Chinese medicine, but also a major source of income for numerous Himalayan residents. Here, taking advantage of rapid advances in single-molecule sequencing, we assembled a highly contiguous genome assembly of O. sinensis. The assembly of 23 contigs was ∼110.8 Mb with a N50 length of 18.2 Mb. We used RNA-seq and homologous protein sequences to identify 8,916 protein-coding genes in the IOZ07 assembly. Moreover, 63 secondary metabolite gene clusters were identified in the improved assembly. The improved assembly and genome features described in this study will further inform the evolutionary study and resource utilization of Chinese cordyceps.