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The medicinal orchid genus Dendrobium belonging to the Orchidaceae family is a huge genus comprising about 800-1,500 species. To better illustrate the species status in the genus Dendrobium, a comparative analysis of 33 available chloroplast genomes retrieved from NCBI RefSeq database was compared with that of the first complete chloroplast genome of D. nobile from north-east India based on next-generation sequencing methods (Illumina HiSeq 2500-PE150). Our results provide comparative chloroplast genomic information for taxonomical identification, alignment-free phylogenomic inference and other statistical features of Dendrobium plastomes, which can also provide valuable information on their mutational events and sequence divergence. [ABSTRACT FROM AUTHOR]

Nepenthes khasiana Hook f., is the only representative member of the genus Nepenthes found in India. The species has great ethnomedicinal importance in addition to its ornamental value. It is used for treatment of several diseases by the local people of Meghalaya. The plant is also characterized by synthesis of bioactive compound called naphthoquinones which is a potent antifungal agent. However, it has become threatened in its natural habitat due to several anthropogenic activities. N. khasiana has been included in the Appendix-I of CITES (Convention on International Trade in Endangered Species of Wild Fauna and Flora). The potential for long term survival of this species in the wild is uncertain, but will surely depend upon the continuing efforts of the local communities to preserve its habitats. The present review highlights the significance of the species and also elucidates the recent findings on the species on various aspects. [ABSTRACT FROM AUTHOR]

Malaxis wallichii (Lindl.) Deb, a small, perennial, monopodial, terrestrial orchid, is endemic to tropical Himalayas at an altitude of 1200-2000 m asl. The pseudobulbs are important ingredients of century old drug 'Ashtavarga' and a polyherbal energetic tonic 'Chyavanprash'. An efficient genetically stable in vitro propagation protocol using transverse thin cell layer culture system was established for M. wallichii. In the present report, meta-topolin alone proved to be three times more beneficial compared to other routinely used cytokinins in inducing highest number of shoot buds, plant height and growth of regenerated shoots. The highest regeneration frequency (89%) along with maximum number of adventitious shoots per explant (22.5 ± 0.6) was observed in MS medium supplemented with 1.0 mg/l meta-topolin and 0.5 mg/l α-naphthalene acetic acid. Highest rooting frequency with highest number of roots (8.66 ± 0.3) was achieved in half-strength MS medium fortified with 1.0 mg/l indole acetic acid. Clonal stability of in vitro-derived plantlets was evaluated and compared to donor plant using intron splice junction (ISJ) markers and flow cytometry. ISJ markers revealed 4.76% clonal variability indicating high degree of genetic stability amongst the in vitro-derived regenerants. The nuclear DNA content of M. wallichii (2n) was found to be 2C = 2.760 ± 0.02 pg and therefore, 1349.64 Mbp (1C). Flow cytometry analysis of actively growing young and mature leaves from donor as well as in vitro-derived plantlets revealed presence of three peaks corresponding to 2C, 4C and 8C, while 2C was the most abundant. In the present investigation, there was no significant difference in the 2C DNA content between the mother and in vitro-derived plants; however, the frequency of endopolyploid cells varied in young and adult plants. An increased HO content as well as lipid peroxidation activities were observed during early stages of acclimatization which declined afterwards. The enhanced activities of antioxidant enzymes like superoxide dismutase, catalase, ascorbate peroxidase and glutathione reductase in acclimatized plantlets as compared to in vitro-grown ones revealed their active involvement in growth and development against oxidative stress under external adverse environment. [ABSTRACT FROM AUTHOR]

An efficient in vitro regeneration protocol has been developed from shoot tips of Drosera burmannii Vahl., a carnivorous plant of north-east India. Various plant growth regulators were used to study their efficacy in the induction of multiple shoots and roots. Of the various treatments, the maximum number of shoots (28.8 ± 1.5) and roots (9.7 ± 0.6) was observed in one-fourth strength standard medium (MS with 50 mg/l citric acid and 10 mg/l ascorbic acid) supplemented with 4 mg/l 6-benzylaminopurine (BAP) and 4 mg/l α-naphthalene acetic acid (NAA) followed by 26.8 ± 1.4 shoots in one-fourth strength SM fortified with 4 mg/l kinetin (KN) and 4 mg/l NAA. The well-developed plantlets with shoots and roots were potted in small plastic glasses filled with a mixture of sand and farmyard manure (3:1); these plantlets when transferred to a glasshouse for hardening and acclimatization showed 90% survival. [ABSTRACT FROM AUTHOR]

Carnivorous plants are among the curiosities of nature being different from the normal plants in their mode of nutrition. These plants have fascinated several researchers for centuries. They are also characterized by synthesis of bioactive compounds which are used as a mechanism for self defense. These compounds possess a broad spectrum of biological activities such as antiparasitic, antibacterial, insecticidal, fungicidal, anti-inflammatory, antipyretic, antiproliferative activities. Although, several antimicrobial drugs have been introduced during the recent decades, the problems of microbial infections resistant to synthetic pesticides still exist which necessitate the introduction of novel antimicrobial agents with additional modes of actions than the currently available therapeutic agents. Naphthoquinones are one of the most studied bioactive compounds which have been reported to inhibit the growth of proliferative cells and microbes. Efforts have been made to induce the biosynthesis of naphthoquinone in different species of carnivorous plants. It has been demonstrated that the accumulation of naphthoquinones in carnivorous plants was increased by injecting chitin into the plant tissues. Also, their biosynthesis could be enhanced by the incorporation of elicitors in in vitro cultures of plants. In the present review, we discuss the applications of naphthoquinones and its biosynthesis in carnivorous plants. [ABSTRACT FROM AUTHOR]

Intercontinental dislocations between tropical regions harboring two-thirds of the flowering plants have always drawn attention from taxonomists and biogeographers. One such family belonging to angiosperms is Orchidaceae with an herbaceous habit and high species diversity in the tropics. Here, we investigate the evolutionary and biogeographical history of the genus Cymbidium, which represents a monophyletic subfamily (Epidendroideae) of the orchids and comprises 50 odd species that are distinctly distributed in tropical to temperate regions. Much is not known about correlations among the level of CAM activity (one of the photosynthetic pathways often regarded as an adaptation to water stress in land plants), habitat, life forms, and phylogenetic relationships of orchids from an evolutionary perspective. A relatively well-resolved and highly supported phylogeny for Cymbidium orchids is reconstructed based on sequence analysis of ITS2 and matK regions from the chloroplast DNA available in public repositories viz. GenBank at NCBI. This study examines a genus level analysis by integrating different molecular matrices to existing fossil data on orchids in a molecular Bayesian relaxed clock employed in BEAST and assessed divergence times for the genus Cymbidium with a focus on evolutionary history of photosynthetic characters. Our study has enabled age estimations (45Ma) as well as ancestral area reconstruction for the genus Cymbidium using BEAST by addition of previously analyzed two internal calibration points. [ABSTRACT FROM AUTHOR]

Nardostachys jatamansi (D. Don) DC., a small, perennial, rhizomatous herb of immense medicinal importance since ancient times, is restricted to specialized habitats of alpine Himalayas ranging from 3000 to 5200 m asl. The species has been recently listed as critically endangered under IUCN Red list of threatened species due to over exploitation of its rhizomes for medicinal uses, habitat degradation, trade and other biotic and anthropogenic interferences. An efficient protocol using both indirect and direct shoot organogenesis has been optimized for N. jatamansi. Best callusing was achieved from the cut ends of leaf and petiole explants within 15 days of culture in MS medium supplemented with 1.5 mg/l α-naphthalene acetic acid and 1.0 mg/l meta-Topolin. Culturing the explants at low temperature (13 ± 1 °C) resulted in better callus growth, shoot regeneration, hyperhydricity control and improvement in photosynthetic pigment content in regenerated shoots. Also, direct organogenesis from shoot tip and petiole explants was achieved in MS medium containing 1.0 mg/l meta-Topolin. Optimum rooting was achieved in the same medium supplemented with 1.0 mg/l indole acetic acid wherein averages of 4.52 roots/shoot were induced. Genetic stability of in vitro-derived plantlets was assessed and compared to mother plant using molecular markers and flow cytometry. Intron Splice Junction (ISJ) and Start Codon Targeted polymorphism (SCoT) marker based profiling revealed uniform banding profile in case of direct shoot organogenesis (DSO)-derived plants while callus mediated organogenesis (CMO)-derived plants showed slight variations as compared to mother plant. The genome size of N. jatamansi was found to be 2C = 1.40 ± 0.01 pg and therefore 684.6 Mbp (1C). Although organogenic calli showed mixoploidy but no major phenotypic and genetic rearrangements were detected by flow cytometry in callus-derived plants. Significantly higher antioxidant activity was observed in callus-derived plants as compared to mother and DSO-derived plants. Plant parts, regeneration pathways and various solvent systems greatly affected the yields of total phenolics, flavonoids, alkaloids, tannins contents present in the in vitro raised plantlets. [ABSTRACT FROM AUTHOR]

Cell suspension cultures were initiated from hypocotyl derived callus to induce capsaicin biosynthesis in suspension cultures of Naga King Chili ( Capsicum chinense Jacq.). Efficient capsaicin production with high growth index (GI) was obtained by exposing cells to salicylic acid (SA) and calcium channel modulators in suspension cultures. The time course of capsaicin formation is related to the cell growth profile in a batch culture. Cells cultivated in the standard medium (SM) initially showed low level of capsaicin yield during active growth. When the cells approached stationary phase, cell growth and cell viability decreased whereas capsaicin production increased continuously. In the fed-batch cultures, the highest capsaicin yield (567.4 ± 8.1 μgg fresh weight) (f.wt) was obtained by feeding the cells with 1 mM SA. However, SA feeding during cultivation repressed the cell growth. Enhanced cell growth (3.1 ± 0.1 GI/culture) and capsaicin yield (534 ± 7.8 μggf.wt) were obtained when the cells were fed with calcium ionophore A23187 (0.5 mM) on day 25 as compared to the control. Addition of the calcium channel blocker verapamil hydrochloride (100 mM) inhibited cell growth and capsaicin production in Naga King Chili suspension cell cultures. [ABSTRACT FROM AUTHOR]

The escalating loss of biological diversity throughout the world has become a major concern for the conservation biologists. Like other threatened plant species, the natural populations of the orchids are also severely threatened. Dendrobium thyrsiflorum is one such representative of the family Orchidaceae whose natural populations are getting destroyed at an alarming rate and deserves special conservation attention. Both direct shoot organogenesis (DSO) and indirect shoot organogensesis (ISO) pathways were experimented and the highest regeneration frequency for DSO and ISO pathways were found to be 86.2 and 96 % respectively. The regenerated shoots were best rooted in half-strength MS medium supplemented with 1 mg/l indole butyric acid (IBA) and 0.5 mg/l phloroglucinol. The genetic stability of the acclimatized plants derived from ISO and DSO pathways was assessed using Inter Simple Sequence Repeats (ISSR) and Start Codon Targeted (SCoT) molecular markers. SCoT proved to be a superior marker over ISSR in detecting clonal variability. The phytochemical analysis of the micropropagated plants also revealed a comprehensively higher yield of various secondary metabolites with significantly higher antioxidant potentials in both ISO- and DSO-derived plants over the mother plant. However, the ISO-derived plants were more phytochemically enriched compared to the DSO-plants. The rapid multiplication rate, higher genetic stability and secondary metabolite production ensures the utility of this micropropagation method for D. thyrsiflorum in the ex situ conservation and commercial exploitation of other important orchid species. [ABSTRACT FROM AUTHOR]

Rapid clonal propagation of selected genotypes has been one of the most extensively exploited approaches of biotechnology. However, inclusion of somaclonal variations in tissue-culture-derived plants results in the production of undesirable plant off-types which limits its applications in tissue culture industry. Therefore, the most critical concern has been the maintenance of genetic uniformity of micropropagated plants. Assessment of genetic fidelity in tissue-culture-raised plants of three consecutive regenerations of Nepenthes khasiana has been successfully carried out using chromosome counts and heterochromatin distribution pattern wherein changes in the number of chromosomes and the distribution of AT and GC base pairs were recorded. The cells studied in the plantlets of the first regeneration (23.33 %) showed deviant number of chromosome which was increased to 33.33 % and 40 % in the plantlets of the second and the third regenerations, respectively. Also, 4′,6-diamidino-2-phenylindole (DAPI) and chromomycin A3 (CMA) binding sites, on an average of 5.74 ± 0.47 and 5.00 ± 0.30, were observed in the plantlets of the first regeneration. Subsequently, DAPI binding sites were increased to 6.61 ± 0.39 and 6.74 ± 0.57 in the plantlets of the second and the third regenerations, respectively, with a corresponding decrease in the CMA binding sites (4.63 ± 0.45 and 4.16 ± 0.47 CMA sites in the plantlets of the second and the third regenerations, respectively). The study reveals an increase in cytological variations in the morphologically similar micropropagated plants of N. khasiana with the subsequent regenerations which further necessitate the determination of genetic integrity of micropropagated plants. [ABSTRACT FROM AUTHOR]

One of the main characteristics of chili pepper fruit is its pungent taste due to the presence of a group of compounds known as capsaicinoids. Capsaicinoids biosynthesis in chili plant is defined by two pathways: phenylpropanoid, which determines phenolic structure; and fatty acid metabolism, which determines the molecule's fatty acids. The potency of chili pepper cells, tissue and organ to biosynthesize capsaicinoids in in vitro cultures have been a subject of intensive research. Recent findings demonstrated that manipulation of culture strategies viz., immobilization of cells, precursors feeding, cell selection, elicitors treatments, osmotic stress, influence of calcium channel modulators, nutrient stress, pH stress etc. to enhance the accumulation of capsaicinoids in in vitro cultures of chili pepper have resulted in significant increase. However, the levels of capsaicinoids accumulation in cell cultures have never reached the levels in the fruits and require more efficient strategies to enhance the capsaicinoids biosynthetic activity. The present review highlights a comprehensive overview of capsaicinoids biosynthesis, its clinical applications and consolidated results of studies on biotechnological advances in chili pepper research leading to the biosynthesis of capsaicinoids in in vitro cultures. [ABSTRACT FROM AUTHOR]

A protocol for induction of direct somatic embryogenesis and subsequent plant regeneration for the medicinally important and endangered plant Paris polyphylla Sm. has been developed for the first time. Immature zygotic embryos (IZEs) were cultured on different media namely Gamborg (B5), ½ B5, Murashige and Skoog (MS), ½ MS, Chu et al. (N6), ½ N6, Schenk and Hildebrandt (SH) and ½ SH. Highest frequency of somatic embryogenesis (32.6 %) and mean number of somatic embryos (SEs) per explant (28.7 ± 1.7) were obtained on ½ MS medium directly without an intermediate callus phase. The frequency of SE induction was significantly increased to 40.7 % when ½ MS medium was solidified with gelrite compared to agar (32.6 %). Secondary somatic embryos (SSEs) appeared on the primary SEs in a repetitive way on plant growth regulator-free ½ MS medium but with a gradual decrease in embryogenic potential during subsequent subcultures. Plasmolyzing pre-treatment of SSEs with 1.0 M mannitol for 12 h effectively maintains its embryogenic capacity. Primary embryos at the elongated dimpled and early cotyledonary stage displayed the highest embryo forming capacity of 26.94 and 27.87, respectively. High frequency of SE germination (94.0 %) occurred on ½ MS medium with 0.5 mg/l gibberellic acid. Highest percentage of seedling to plantlet conversion was observed in the medium supplemented with 0.05 mg/l 6-benzylaminopurine and 0.1 mg/l α-naphthalene acetic acid. Regenerated plants displayed morphological characteristics similar to that of the wild plants. Flow cytometry analysis showed ploidy stability of the regenerated plants. [ABSTRACT FROM AUTHOR]

Proteins and activities of antioxidant enzymes, including polyphenol oxidase (PPO), peroxidase (POX) and catalase (CAT), were evaluated in Dendrobium hookerianum during different stages of development of both protocorms and protocorm-like bodies (PLB) derived from seeds and axillary buds, respectively. The changes in the protein contents and the enzyme activities along with their isozyme patterns were compared between these two culture systems. It was found that the protein contents and the enzyme activities increased steadily over time during different stages of development in both the systems. Protein contents (4.57 mg/g fresh wt.) and activities of POX (21.9 U/mg protein), CAT (9.86 U/mg protein) were observed to be higher in PLB system as compared to protocorm system at stage IV of development. However, although the PPO activity increased gradually till the third stage of development, a decline was observed at stage IV wherein the activity was recorded to be more or less same in both the systems. Also, few proteins (~61, 50, 46, 32, 25, 16, 6 kDa) and new isozymes (POX 7, 8; CAT 2) were expressed only in PLB system of development. In general, high protein content and enzyme activities were detected in PLB system as compared to protocorm system. The results of the present study indicate that few proteins and isozymes could be regarded as specific biochemical markers for different stages of development of this medicinally important orchid. [ABSTRACT FROM AUTHOR]

Manipulation of culture strategies was adopted to study the influence of nutrient stress, pH stress and precursor feeding on the biosynthesis of capsaicin in suspension and immobilized cell cultures of C. chinense. Cells cultured in the absence of one of the four nutrients (ammonium and potassium nitrate for nitrate and potassium stress, potassium dihydrogen orthophosphate for phosphorus stress, and sucrose for sugar stress) influenced the accumulation of capsaicin. Among the stress factors studied, nitrate stress showed maximal capsaicin production on day 20 (505.9 ± 2.8 μg g f.wt) in immobilized cell, whereas in suspension cultures the maximum accumulation (345.5 ± 2.9 μg g f.wt) was obtained on day 10. Different pH affected capsaicin accumulation; enhanced accumulation of capsaicin (261.6 ± 3.4 μg g f.wt) was observed in suspension cultures at pH 6 on day 15, whereas in case of immobilized cultures the highest capsaicin content (433.3 ± 3.3 μg g f.wt) was obtained at pH 5 on day 10. Addition of capsaicin precursors and intermediates significantly enhanced the biosynthesis of capsaicin, incorporation of vanillin at 100 μM in both suspension and immobilized cell cultures resulted in maximum capsaicin content with 499.1 ± 5.5 μg g f.wt on day 20 and 1,315.3 ± 10 μg g f.wt on day 10, respectively. Among the different culture strategies adopted to enhance capsaicin biosynthesis in cell cultures of C. chinense, cells fed with vanillin resulted in the maximum capsaicin accumulation. The rate of capsaicin production was significantly higher in immobilized cells as compared to freely suspended cells. [ABSTRACT FROM AUTHOR]

Capsicum species commonly known as Chili peppers are economically important group of plants belonging to the Solanaceae family. Of the 38 species reported, only six species namely, Capsicum annuum, C. assamicum, C. baccatum, C. frutescence, C. chinense and C. pubescens are cultivated. They are very important component of the human being as peppers are used as vegetables, spices, and a coloring agent and for medicinal purposes. Based on pungency trait which is due to the presence of a group of compounds known as capsaicinoids, cultivated capsicums are classified into sweet peppers and hot peppers. Although conventional breeding and classical genetic analysis were successful in estimating the number of genes for economically important traits governed by few major genes and their incorporation in the breeding programme, the advent of molecular markers and recently developed next generation sequencing technologies supplemented greatly in dissecting the genetic and molecular basis of economically important traits in the capsicum genome for applied research. Here in this review, we tried to highlight the use of molecular markers, comparative mapping and advanced genomics technologies and their integrated use in the translational research of cultivated Capsicums. [ABSTRACT FROM AUTHOR]

Different dilutions, that is, 25, 50, 75, and 100%, of tannery effluent (TE) were chosen for the present study to assess the phytotoxic effects on Vigna mungo L. For amelioration purposes, different levels and combinations of iron and zinc were supplied to the plants along with 50% TE that is chosen on the basis of prior test under Petri dish culture. Cytotoxic and biochemical analysis and plant tolerance index (PTI) of plant were observed. Mitotic index deceased with increase in effluent concentration whereas abnormality % was increased. The pigments (chlorophyll a, total, and carotenoids) were decreased with increasing treatment levels of TE at both growth stages. However, carotenoid content increased significantly at all dilution levels of TE after first growth stage. Chlorophyll b was increased significantly after 35 days of growth but decreased after 70 days. Theprotein contentswere also significantly decreased with increase in all TE treatments and increased significantly in zinc recovery treatments. Activities of catalase and peroxidase enzymeswere significantly affected and increased significantly with effluent treatments. PTI showed an enhanced tolerance capacity of plant with treatment of iron and zinc. A negative correlation was found (r = -0.97) between plant height and different dilutions of effluent whereas it was positively correlated (r = 0.95) with iron and zinc treatments. The study represents the ameliorative effect of iron and zinc for phytotoxic damage in V. mungo caused by tannery effluent. [ABSTRACT FROM AUTHOR]

Establishment of symbiotic association at the appropriate developmental stage helped maintain continued growth which is vital for the long-term ex vitro survival of the orchid. In the present study, symbiotic association was carried out using different developmental stages of Dendrobium chrysanthum and pathogenic Rhizoctonia isolates (obtained from orchids and non-orchid hosts) in different culture media. Isolate 2162 supported highest symbiotic germination on OMA-S (oat meal agar medium without nutrients + sucrose), whereas, stable symbiotic association with plantlets was obtained with isolate 4634 on OMA-NC (oat meal agar medium + cellulose). Isolate Dc-2S2 obtained from the host plant did not promote seed germination nor did it form association with protocorms or plantlets. This study, for the first time identifies a combination of compatible fungal isolate, suitable culture medium, and appropriate developmental stage at which symbiotic association in vitro can be deemed successful for the medicinally important orchid, D. chrysanthum. [ABSTRACT FROM AUTHOR]

An efficient in vitro protocol for large-scale multiplication of Nepenthes khasiana, a threatened insectivorous plant of India, has been developed from nodal stem segments. The highest shoot proliferation of 19.16 ± 0.23 shoots/explant was recorded in half-strength Murashige and Skoog (MS) medium supplemented with 2.5 mg/l kinetin, 2.0 mg/l 6-benzyl aminopurine, 3 % sucrose and 0.8 % agar. The best rooting was achieved in half-strength MS medium supplemented with 2.0 mg/l α-naphthalene acetic acid with an average of 9.04 ± 0.46 roots/shoot. The plantlets were successfully transferred to the greenhouse with survival rate of 92 %, exhibiting normal development. Cytological and random amplified polymorphic DNA (RAPD) analyses were carried out to assess the genetic integrity of the regenerated plantlets. Cytological analysis revealed no change in chromosome number with cells studied showing 2 n = 80. Of the 80 primers screened for RAPD analysis, 14 primers resulted in clear and scorable bands. A total of 72 amplification products were obtained out of which only 4.1 % bands were polymorphic. Cluster analysis of the RAPD profile revealed an average similarity coefficient ranging from 0.98 to 1.0, thus suggesting genetic stability in the micropropagated plants of N. khasiana. [ABSTRACT FROM AUTHOR]

The role of mycorrhizal fungi in overcoming nutrient limitation to plant growth by enhancing nutrient acquisition, especially phosphorus (P) and nitrogen (N), is well documented. However, in orchids, the importance of mycorrhizal fungi in nutrient acquisition is not as extensively studied as in other plants. Therefore, an in vitro culture system to study the effects of mycorrhizal association on P and N metabolizing enzymes, viz., acid phosphatase, alkaline phosphatase, nitrate reductase (NR), nitrite reductase (NiR) and glutamine synthetase (GS) in Dendrobium chrysanthum was developed. After 90 days of mycorrhizal treatment, activities of acid phosphatase, alkaline phosphatase, NR, NiR and GS were higher in mycorrhizal plantlets than in control plantlets. The hardened plantlets that were initially treated with mycorrhiza under in vitro conditions also showed higher activities of the enzymes investigated. These mycorrhizal plantlets showed higher survival (96.33 %), shoot length (3.7 cm) and shoot fresh weight (0.359 g) as compared to control after 120 days of hardening. The results presented in this study suggest that mycorrhizal association might have increased the assimilation of P and N in D. chrysanthum plantlets, indicating the importance of mycorrhiza in orchids. [ABSTRACT FROM AUTHOR]

A successful cryopreservation protocol for the long-term conservation of protocorms of two threatened and vulnerable orchids, Cymbidium eburneum Lindl. and Cymbidium hookerianum Rchb. f., was developed using encapsulation-dehydration. Protocorms were osmoprotected in liquid Murashige and Skoog medium (MS) containing 0.7 M sucrose for 20 h at 25 ± 2°C on a rotary shaker, and incorporated into an encapsulation matrix [consisting of 3% ( w/ v) sodium alginate and 100 mM CaCl]. The encapsulated protocorms, which were desiccated in a laminar airflow cabinet for 6 h, were able to withstand cryostorage in liquid nitrogen. Maximum regeneration into complete plantlets (72% for C. eburneum and 70% for C. hookerianum) of the cryostored, encapsulated protocorms was obtained using MS medium containing 3% sucrose and 0.8% agar. Using this protocol of cryopreservation, long-term preservation for ex situ conservation of these two threatened orchids can be accomplished. [ABSTRACT FROM AUTHOR]

Synthetic seed technology is an exciting and rapidly growing area of research as deals with conservation and storage of rare, endangered and desirable genotypes along with its easy handling and transportation. As propagation of many ornamental and medicinally important plant species is labour intensive, application of different growth retardants and osmotica in simple artificial seed system would dramatically reduce labour requirement by storing the germplasm in vitro. Moreover, the primary aim of developing in vitro storage methods is to reduce the frequent demands of subculturing and preserving the unique genetic constituent of the germplasm. Dendrobium nobile is a pharmaceutically important orchid mostly used in the Chinese herbal drug industry for its medicinal property. Commercial exploitation of this species has considerably depleted their population in wild. Hence, for conserving this valuable germplasm, short term in vitro storage of Protocorm-Like Bodies (PLBs) of D. nobile was carried out using different osmotica (sucrose and mannitol). It was observed that incorporation of low sucrose and mannitol (3 and 5 %) in the encapsulating matrix showed almost similar results with that of control. In all these cases, more than half of PLBs burst out from the matrix thus making these concentrations of sucrose and mannitol along with control not suitable for storage studies. However, with the increase in concentration to 7.5 and 12.5 % in the encapsulating matrix, no outburst of encapsulated PLBs was recorded till 60 days of storage; hence it can be concluded that these concentrations play an important role in minimizing the growth of PLBs during storage condition. [ABSTRACT FROM AUTHOR]

Thirty-five inter-simple sequence repeat (ISSR) markers were used to analyze the genetic variation in Cymbidium spp. High number of polymorphic bands (217) with overall 90 % of polymorphism at inter-specific level was observed. Cumulative genetic similarity ranged from 0.40-0.93 with an average value of 66 % among the species. At intra-specific level, average polymorphism detected, ranged from 29.8 to 69.9 % within the five species of Cymbidium. All the species were apparently endowed with low genetic variation at intra-specific level compared to inter-specific level. UPGMA clustering evidently distinguished the representatives of C. aloifolium and C. tigrinum which may be linked to entirely different climatic conditions in which they grow, besides their discrete morphological characteristics. Nine ISSR primers revealed 11 unique species-specific banding patterns belonging to three Cymbidiums, which can further developed as SCAR markers. Thus, present investigation provides valuable baseline data of genetic variation in five species of Cymbidium and addresses the conservation concerns of this horticulturally important orchid. [ABSTRACT FROM AUTHOR]

A successful report on the in vitro propagation of Homalomena aromatica via rhizome axillary bud multiplication is presented. Rhizome bud explants were cultured on Murashige and Skoog medium supplemented with various concentrations of cytokinins to induce multiple shoot formation for micropropagation. The highest number of shoots was achieved in MS medium supplemented with 2.0 mg l 6-benzylaminopurine. The regenerated shoots rooted most efficiently on half-strength MS medium supplemented with 0.5 mg l α-naphthalene acetic acid. The regenerated plantlets showed no morphological differences from the parent plant. This protocol takes approximately 6 months to reach the acclimatization stage from the initiation stage and facilitates commercial and rapid propagation of H. aromatica. [ABSTRACT FROM AUTHOR]

A comprehensive reassessment of IndianNymphaea(Nymphaeaceae) based on morphology, RAPD and nucleotide sequence data of the ITS,trnKintron,matKandrbcLgene was conducted. Although considerable morphological variations have been reported, pollen colour and rhizome shape are two characters which have not been mentioned in previous studies. The transformation from yellow to white coloured pollen may have evolved independently and is probably associated with a strong selective pressure acted upon by the animal pollinators on white pollen ofN. pubescensandN. rubra. Rhizome shape could easily discriminate among subgenera; the difference in shape (triangular, globular and cylindrical) probably highlights the success in their distribution. Our studies using RAPD revealed high genetic variability among individuals ofN. caerulea. This may be attributed to the breeding system followed, which could be an outcrossing species. But the lack of genetic diversity inN. tetragonais probably due to founder events, whereby a solitary founder individual from China could have resulted in the establishment of this single population of ∼25–30 individuals with no detectable variations. Molecular cloning of the ITS region ofN. rubra, necessitated by the presence of additional signals in the sequencing chromatogram, confirmed the origin of this plant taxon through hybridization involvingN. lotusandN. pubescensas the parental species. [ABSTRACT FROM AUTHOR]

The population of many splendid orchids is reducing from their natural habitats at an alarming rate and their conservation is becoming a matter of global concern. Asymbiotic seed germination has been applied for ex situ conservation of rare, endangered and threatened orchid taxa and could provide rapid means their multiplication. In the present study reported here, seeds of an epiphytic and rare orchid, Cymbidium eburneum were germinated asymbiotically in different basal media viz., Murashige and Skoog (MS), Knudson C, Mitra et al. (Mitra), Gamborg et al. (B) and Nitsch. The highest germination rate was observed in Mitra medium, whereas the development of the protocorms was found to be best in MS medium. Effects of growth regulators viz., indole-3 acetic acid (IAA), α-naphthalene acetic acid (NAA), 2,4-dichlorophenoxyacetic acid (2,4- d), thidiazuron (TDZ), 6-benzyl aminopurine (BAP) and kinetin (Kn) both singly and in combination incorporated in the MS medium were studied on growth and development of seedlings. It was observed that MS medium nourished with 15 μM each of BAP and NAA in combination was found to enhance shoot number and length, and root number and length in the seedlings. The rooted seedlings were successfully acclimatized . [ABSTRACT FROM AUTHOR]

Understanding the genetic resources and diversity is very important for the breeding programs and improvement of several economically important orchids like Cymbidium. Karyomorphological studies have been carried out on seven Cymbidium species, C. aloifolium (Linnaeus, 1753), C. devonianum Paxton, 1843, C. elegans Lindley, 1828, C. iridioides D. Don, 1825, C. lowianum Rchb. f.,1877, C. tigrinum Parish ex Hook. f., 1864, and C. tracyanum L. Castle,1890, most of them endangered/threatened in their natural habitat. As reported earlier, the somatic chromosome number (2n = 40) has been observed in all the seven species. Distinct inter-specific variation was recorded in the arm ratio of few homologous pairs in the complements. Symmetrical or almost symmetrical karyotypes were prevalent; however significant asymmetry was reported in C. iridioides and C. tracyanum. The significance of karyotypic variation in speciation of the genus Cymbidium has been discussed. This study provides useful chromosome landmarks and evidence about genome evolution, heteromorphic chromosomes based heterozygosity, basic chromosome number and ploidy level in the genus Cymbidium. [ABSTRACT FROM AUTHOR]

The influence of osmotic stress on capsaicin production was investigated in cell suspension cultures of Capsicum chinense Jacq.cv. Naga King Chili, a chili species native to Northeastern India. The sterilized seeds were germinated in Murashige and Skoog medium. Two-week-old hypocotyls were excised from in vitro germinated seedlings and implanted in MS medium containing 2, 4-dichlorophenoxyacetic acid (2 mg/l), and Kinetin (0.5 mg/l) for callus induction. Capsaicin production in the suspension cultures was significantly affected using sucrose, mannitol, and NaCl in the medium. Stoichiometric analysis with different combinations of sucrose and non-sugar osmotic agent (NaCl) showed that osmotic stress was an important factor for enhancing capsaicin production in cell suspension cultures of C. chinense. The capsaicin content of 1,644.1 μg g f.wt was recorded on day 15 in cultures grown in MS medium containing 87.64 mM sucrose in combination with 40 mM NaCl. However, osmotic stress treatment at 160 mM NaCl with sucrose resulted in lowering capsaicin accumulation and separation of cell wall from their cytoplasm, under microscopic observation. [ABSTRACT FROM AUTHOR]

Random Amplified Polymorphic DNA (RAPD) markers were utilized for determination of diversity within and among the three populations of Nepenthes khasiana Hook f., a threatened insectivorous plant of Meghalaya (India). A total of 90 bands were generated from 10 random amplification polymorphic DNA (RAPD) primers of which 71 were found to be polymorphic (78.89%). Nei's gene diversity ( h) ranged between 0.124-0.201 with overall diversity of 0.228 while Shannon's information index I) values recorded between 0.187-0.308 with an average value of 0.352. The values of gene flow ( Nm = 1.284) and the diversity among populations (0.280) recorded demonstrates higher genetic variation within the population. AMOVA analysis revealed a low level of genetic variation (21.96%) among the populations. This study indicates that some variation still exists within and between the existing populations of N. khasiana, thus, these populations could provide materials for re-establishing of this important rare and threatened species. [ABSTRACT FROM AUTHOR]

Karyo-morphological studies have been carried out in two critically rare, endangered and endemic rhizomatous, horticulturally important species of genus Mantisia (Zingiberaceae) from North-East India. The somatic chromosome number (2n = 20) has been recorded in both M. spathulata and M. wengeri with distinct inter-specific variation in the arm ratio of homologous pairs in the complements. All the chromosomes were sub-metacentric in nature (20 L) and more or less symmetrical karyotypes in both the species indicating stabilized genomic pattern. In M. wengeri two chromosome pairs exhibited heteromorphy. Nucleolar organizing region (NOR) and meta-, telo-, and sub-telocentric chromosomes were absolutely lacking. Chromosome class based on total length of each chromosome as well as its designation through centromeric position was determined. This karyo-morpological study provides valuable baseline genetic data for clearly distinguishing Mantisia ( x = 10) from its phenotypically very similar allied genus Globba L. ( x = 8). Thus, present investigation provides valuable insight on karyotypic variation, speciation and evolution of the genus Mantisia and also addresses the conservation concerns. [ABSTRACT FROM AUTHOR]

True endomitosis in the anther tapetum of the three Cymbidium species viz. C. aloifolium, C. devonianum and C. tigrinum is described. In these Cymbidiums, most tapetal cells go through endomitosis instead of either normal mitosis or so called 'inhibited' mitosis. The nuclear membrane does not disappear, but during metaphase the chromosomes are considerably condensed far more than in normal mitosis, and nucleolus persists throughout the endomitotic cycle. Endomitosis may not be unusual to the tapetal cells of these peculiar Cymbidiums but may have a wider application and explain many of the cytological phenomena occurring in the tapetal cells of other plants. Further, physical localization of 45S rRNA genes during endomitotic divisions may help to confirm their atypical activities as well as provide insight into the course and degree of tapetum polyploidization. Analysis of DNA-methylation levels is also recommended for understanding the role of nucleolus for spindle formation during endomitosis. [ABSTRACT FROM AUTHOR]

Background: The Nymphaeales (waterlilly and relatives) lineage has diverged as the second branch of basal angiosperms and comprises of two families: Cabombaceae and Nymphaceae. The classification of Nymphaeales and phylogeny within the flowering plants are quite intriguing as several systems (Thorne system, Dahlgren system, Cronquist system, Takhtajan system and APG III system (Angiosperm Phylogeny Group III system) have attempted to redefine the Nymphaeales taxonomy. There have been also fossil records consisting especially of seeds, pollen, stems, leaves and flowers as early as the lower Cretaceous. Here we present an in silico study of the order Nymphaeales taking maturaseK (matK) and internal transcribed spacer (ITS2) as biomarkers for phylogeny reconstruction (using character-based methods and Bayesian approach) and identification of motifs for DNA barcoding. Results: The Maximum Likelihood (ML) and Bayesian approach yielded congruent fully resolved and wellsupported trees using a concatenated (ITS2+ matK) supermatrix aligned dataset. The taxon sampling corroborates the monophyly of Cabombaceae. Nuphar emerges as a monophyletic clade in the family Nymphaeaceae while there are slight discrepancies in the monophyletic nature of the genera Nymphaea owing to Victoria-Euryale and Ondinea grouping in the same node of Nymphaeaceae. ITS2 secondary structures alignment corroborate the primary sequence analysis. Hydatellaceae emerged as a sister clade to Nymphaeaceae and had a basal lineage amongst the water lilly clades. Species from Cycas and Ginkgo were taken as outgroups and were rooted in the overall tree topology from various methods. Conclusions: MatK genes are fast evolving highly variant regions of plant chloroplast DNA that can serve as potential biomarkers for DNA barcoding and also in generating primers for angiosperms with identification of unique motif regions. We have reported unique genus specific motif regions in the Order Nymphaeles from matK dataset which can be further validated for barcoding and designing of PCR primers. Our analysis using a novel approach of sequence-structure alignment and phylogenetic reconstruction using molecular morphometrics congrue with the current placement of Hydatellaceae within the early-divergent angiosperm order Nymphaeales. The results underscore the fact that more diverse genera, if not fully resolved to be monophyletic, should be represented by all major lineages. [ABSTRACT FROM AUTHOR]

Nymphaea, an aquatic perennial herb with exceptionally beautiful flowers and floating leaves, is well represented globally. Out of ten species reported from India, the internal transcribed spacers (ITS) region of nrDNA was investigated in seven species of Nymphaea viz. N. alba var. rubra, N. caerulea, N. × marliacea, N. nouchali, N. pubescens, N. rubra and N. tetragona. Barring N. pubescens, whereby double peaks detected in the sequencing chromatograms may be due to random mutations occurring in some of the ITS paralogues, the additional signals detected for N. alba var. rubra and N. rubra are probably influenced by recent hybridization and introgression. Our study on sequence characteristics of ITS 1 and ITS 2 revealed high G + C content (ITS 1, 45.5-48.4%; ITS 2, 50.2-51.5%) and sequence divergence. Percentage of sequence divergence based on substitution and substitution plus indels is 44.15 and 57.19, respectively, for the ITS 1; 29.74 and 47.96% were recorded for the ITS 2. Although highly variable, conserved motifs within the ITS 1 and ITS 2 region of Nymphaea were identified and are found to be common throughout the order Nymphaeales. Sequence analysis of the ITS 1 and ITS 2 failed to detect any variation between two morphotypes of N. nouchali, namely N. nouchali JD 06 and N. nouchali JD 07, differing in flower color and found at the same geographical location. However, on comparison with another specimen of N. nouchali found at a different location, they showed considerable variation in nucleotide composition. Complemented by sequence data retrieved from GenBank, phylogenetic tree reconstruction of the genus Nymphaea based on neighbor-joining, maximum parsimony, maximum likelihood and Bayesian inference methods is presented and discussed. [ABSTRACT FROM AUTHOR]

Sustainable utilization of plant genetic resources for food and agriculture has been increasingly discussed at both national and international forums. Besides exploitation, conservation of plant genetic resources has become an integral part of these discussions. Conservation aims at maintaining the diversity of living organisms, their habitat and the interrelationship between organisms and their environment. For achieving such goals, appropriate conservation strategies have to be adopted. Determining the genetic makeup of a particular plant species is of critical importance when planning a suitable conservation strategy. In this study, we sequenced the chloroplast trnK intron, matK and rbcL gene aimed at understanding the rarity of Nymphaea tetragona, a critically rare and endangered plant of India found at only one location. We extended our investigation to other Nymphaea species such as N. nouchali, N. pubescens and N. rubra that are commonly available throughout India. Interestingly, matK gene of N. tetragona revealed high number of non-synonymous substitutions. Molecular evolutionary analysis indicated that three of these sites may be under mild selective pressures. Such adaptive changes at the DNA and protein sequence level of matK gene may have been associated with the colonization of N. tetragona, suggesting that it could have migrated from China. [ABSTRACT FROM AUTHOR]

Clonal propagation is an important biotechnological tool for the conservation of rare, endangered and threatened as well as commercially important plant species of which orchids are an important representative. In the present research, an attempt was made to develop a fast, clonally stable as well as phytochemically enriched regeneration protocol for propagating Dendrobium aphyllum, an important medicinal orchid using transverse thin cell layer (t-TCL) explants. Murashige and Skoog (MS) medium supplemented with 15 µM meta-topolin along with 10 µM TDZ and 10 µM AgNO3 was found to be most conducive for shoot proliferation. After 8 weeks of culture, an average of 39.41 shoots/explant proliferated; till date this is the best shooting frequency for D. aphyllum and one of the best amongst the orchids. The highest rooting frequency of 82.34% was accomplished in half-strength MS medium supplemented with 15 µM IBA. The plantlets were successfully acclimatized in greenhouse exhibiting normal developmental patterns and flowering occurred after 6 months of acclimatization with 3-4 flowers/stalk irrespective of the flowering season. The genetic homogeneity of the micropropagated plants was ascertained using IRAP and ISSR markers. A comparative assessment of the antioxidant activity (mother and micropropagated plant) was performed using DPPH and FRAP assays which was recorded to be higher amongst the in vitro-regenerated plants. A rapid, genetically stable regeneration protocol coupled with comprehensively higher phytochemical yield is reported in the present study on D. aphyllum. The results of this study confirm the use of in vitro propagation protocol for sustainable commercial utilization as well as conservation of medicinal orchid bio-resources within a limited time period. [ABSTRACT FROM AUTHOR]

A correction to the article "Phyto-molecular profiling and assessment of antioxidant activity within micropropagated plants of Dendrobium thyrsiflorum: a threatened, medicinal orchid," by Paromik Bhattacharyya and colleagues, that was published online on July 8, 2015 is presented.