Pratibha Misra

CSIR-National Botanical Research Institute - India · Tissue Culture and Plant Transformation

In the present scenario, remediation of heavy metals (HMs) contaminated soil has become an important work to be done for the well-being of human and their environment. Phytoremediation can be regarded as an excellent method in environmental technologies. The present contemporary research explores the Solanum viarum Dunal function as a potential acc...

Glycosyltransferase (GT) enzymes are the members of a large multigene family in plants that can transfer activated sugar molecules to an extensive range of acceptors, such as sterols and secondary metabolites. This glycosylation of plant metabolites helps in the fortifying defense of the plant against different environmental stress. Sterol glycosyl...

Key message Overexpression of Withania somnifera SGT gene (WssgtL3.1) in transgenic Arabidopsis improves various agronomic and physiological traits and alters conjugated sterol levels to mitigate the effect of salt stress. Abstract Sterols are essential constituents of cell membranes that are involved in several biological functions, including res...

The medicinal plant, Solanum viarum Dunal includes a number of compounds with important pharmacological effects. The effect of temperature (heat and cold) stress on growth, secondary metabolite levels and defense mechanism in two genotypes (prickled and prickleless) of S. viarum was evaluated. The two thermal regimes (4°C for cold and 35°C for heat...

Solanum viarum Dunal, is an important medicinal plant widely used as a source of raw material for the steroidal drug industry. Out of various steroidal precursors, solasodine, an analogue of diosgenin is the most important source of raw material for the synthesis of steroidal drugs. In the present study, growth and phytochemical variations in diffe...

Thalictrum foliolosum is an endemic herb of the temperate Himalayas and eastern China. These plants have been used for past several decades by indigenous people to treatment of fever, gastrointestinal disease, intestinal obstruction, jaundice and eyesight problems. Although a wide diversity of alkaloids and polyphenolic compounds are found in vario...

Solanum is one of the largest genera of the family Solanaceae comprising > 2000 species distributed mostly in the tropical and subtropical regions of Australia, Africa, and some parts of Asia, such as China, India, and Japan. The nutraceutical and pharmaceutical values of the Solanum species are due to the presence of bioactive phyto-constituents s...

Thalictrum foliolosum DC. is an endemic herbaceous plant known for its various medicinal properties. Due to the presence of valuable alkaloids they are uprooted leading to a threat to their numbers and existence. In this study an efficient protocol has been developed for plant regeneration using hypocotyl explant. Hypocotyl explants formed multiple...

Solanum viarum Dunal is an important medicinal plant with a high quantity of steroidal alkaloids used for the synthesis of contraceptives, corticosteroids, and sex hormones. It is also used by Indian tribal people for the treatment of leprosy, toothache, and diabetes. Therefore, to meet the existing needs for this plant, it is necessary to develop...

Thalictrum foliolosum is an endemic herb known for its medicinal properties and used for various clinical applications including ophthalmic, skin disease and dyspepsia. Due to its medicinal properties, the plants are uprooted hence can be prone to extinction. In the present study, a reproducible in vitro propagation protocol has been developed usin...

Growth and production kinetics of three important glycoalkaloids viz. α-solanine, solanidine, and solasodine in two contrasting prickly and prickleless plants of Solanum viarum Dunal were evaluated under in vitro conditions. The prickleless plants showed improved accumulation of total glycoalkaloid content [7.11 and 6.85 mg g−1 dry weight (DW)] and...

Glycyrrhiza glabra L. has become an endangered medicinal plant due to the unabated extraction of glycyrrhizin. Glycyrrhizin is a triterpenoid saponin that is a root centric secondary metabolite having numerous pharmacological properties, such as anti-inflammatory, immunomodulatory, antiallergic, antiulcer, and is found to be effective even against...

Enhancement of secondary metabolites through elicitation in hairy root culture is a very effective method which is broadly used to simulate the stress responses in plants. Elicitors are compounds that induce plants to produce secondary metabolites at elevated levels and reduce the processing time required to achieve high product concentrations. Hai...

Steroidal alkaloids (SAs) are widely synthesized and distributed in plants manifesting as natural produce endowed with potential for medicinal, pesticidal and other high-value usages. Glycosylation of these SAs raises complex and diverse glycosides in plant cells that indeed govern numerous functional aspects. During the glycosylation process of th...

Withania somnifera, commonly known as Ashwagandha an important medicinal plant largely used in Ayurvedic and indigenous medicine for over 3,000 years. Being a medicinal plant, dried powder, crude extract as well as purified metabolies of the plant has shown promising therapeutic properties. Withanolides are the principal metabolites, responsible fo...

Heat is a major environmental stress factor that confines growth, productivity, and metabolism of plants. Plants respond to such unfavorable conditions through changes in their physiological, biochemical and developmental processes. Withania somnifera, an important medicinal plant, grows in hot and dry conditions, however, molecular mechanisms rela...

Sterol glycosyltransferases (SGTs) catalyze the attachment of a carbohydrate moiety to an aglycone sterol accepter molecule at different positions. SGTs are key enzymes for the biosynthesis of many precious natural plant products. SGTs of Withania somnifera (WsSGTs) help in the glycosylation of withanolides, a pharmaceutically important C-28 phytoc...

An ever-increasing demand of uniform plants of commercially valuable plant species needs their clonal propagation on a large scale using different strategies of tissue culture. We have reported a large number of plants which were propagated in vitro using different plant parts. A number of problems were solved before large-scale propagation of thes...

Sterol glycosyltransferases (SGTs) catalyze the attachment of a carbohydrate moiety to an aglycone sterol accepter molecule at different positions. SGTs are key enzymes for the biosynthesis of many precious natural plant products. SGTs of Withania somnifera (WsSGTs) help in the glycosylation of withanolides, a pharmaceutically important C-28 phytoc...

In Withania somnifera, sterol molecules of immense medicinal value are diversified by means of glycosyl-ation. Identifying sterol glycosyltransferases provides an imperative insight of diverse sterol modifications, thereby helping to comprehend the underlying plant mechanisms. In the p r e s e n t s t u d y, o n e o f t h e W. s o m n i f e r a s t...

Sterol glycosyltransferases (SGTs) catalyse transfer of glycon moiety to sterols and their related compounds to produce diverse glyco-conjugates or steryl glycosides with different biological and pharmacological activities. Functional studies of SGTs from Withania somnifera indicated their role in abiotic stresses but details about role under bioti...

Background: Rauwolfia serpentina and Solanum khasianum are well-known medicinally important plants contained important alkaloids in their different parts. Elicitation of these alkaloids is important because of associated pharmaceutical properties. Targeted metabolites were ajmaline and ajmalicine in R. serpentina; solasodine and α-solanine in S. k...

Sterol glycosyltransferases (SGTs) catalyse transfer of glycon moiety to sterols and their related compounds to produce diverse glyco-conjugates or steryl glycosides with different biological and pharmacological activities. Functional studies of SGTs from Withania somnifera indicated their role in abiotic stresses but details about role under bioti...

Key message Overexpression of sterol glycosyltransferase (SGTL1) gene ofWithania somniferashowing its involvement in glycosylation of withanolide that leads to enhanced growth and tolerance to biotic and abiotic stresses. Abstract Withania somnifera is widely used in Ayurvedic medicines for over 3000 years due to its therapeutic properties. It con...

Plants by nature protect themselves by producing secondary metabolites which often have pronounced bioactivities. The presence of these metabolites is responsible for the active interaction of plants with their environment defending themselves against a variety of herbivores and pathogenic microorganisms as well as various kinds of abiotic stresses...

The climbers have very important role as ornamentals (e.g. Bougainvillea, Clematis, Bignonia, Lonicera, floribunda rose, Asparagus racemosus, Gloriosa superba, Trachelospermum jasminoides, Ceropegia spp., etc.) and as medicinal plants (e.g. Gymnema sylvestre, Tinospora cordifolia, Gloriosa superba, Asparagus racemosus, Piper betle, Holostemma ada-k...

Sterol glycosyltransferases regulate the properties of sterols by catalyzing the transfer of carbohydrate molecules to the sterol moiety for the synthesis of steryl glycosides and acyl steryl glycosides. We have analyzed the functional role of TTG15/UGT80B1 gene of Arabidopsis thaliana in freeze/thaw and heat shock stress using T-DNA insertional sg...

Sterol glycosyltransferases (SGTs) belong to family 1 of glycosyltransferases (GTs) are enzymes responsible for synthesis of sterol -glucosides (SGs) in many organism. WsSGTL1 is a SGT of Withania somnifera has been found associated with plasma membranes. However its biological function in W.somnifera is largely unknown. In the present study, we ha...

Welwitschia mirabilis is a phylogenetically important primitive gymnosperm of the monogeneric family Welwitschiaceae. The plant is unique in appearance, having only two leaves throughout its life span. It is a long-lived, slow growing plant and could prove as an excellent experimental material for the study of antiageing gene. Because of non-availa...

Withania somnifera (L.) Dunal, commonly called Indian Ginseng (Ashwagandha), is an important medicinal plant having a number of chemotypes which vary in their regenerative potentiality. In the present study, regenerative potentiality of four chemotypes of W. somnifera was compared using leaf explants. The study correlated that the chemotype (NIMTLI...

Sterol glycosyltransferases (SGTs) are ubiquitous but one of the most diverse group of enzymes of glycosyltransferases family. Members of this family modulate physical and chemical properties of secondary plant products important for various physiological processes. The role of SGTs has been demonstrated in the biosynthesis of pharmaceutically impo...

Glycosylation of sterols, catalysed by sterol glycosyltransferases (SGTs), improves the sterol solubility, chemical stability and compartmentalization, and helps plants to adapt to environmental changes. The SGTs in medicinal plants are of particular interest for their role in the biosynthesis of pharmacologically active substances. WsSGTL1, a SGT...

The commercial importance of linseed (Linum usitatissimum L.) has attracted breeders to increase its seed yield using various breeding approaches. Plant growth regulators (PGRs) have a significant role in enhancing yield and its related traits in linseed. In the present study, two plant growth hormones, auxin and gibberellic acid, were applied indi...

Background: Sterol glycosyltrnasferases (SGT) are enzymes that glycosylate sterols which play important role in plant adaptation to stress and are medicinally important in plants like Withania somnifera. The present study aims to find the role of WsSGTL1 which is a sterol glycosyltransferase from W. somnifera, in plant's adaptation to abiotic stres...

Sterol glycosyltrnasferases (SGT) are enzymes that glycosylate sterols which play important role in plant adaptation to stress and are medicinally important in plants like Withania somnifera. The present study aims to find the role of WsSGTL1 which is a sterol glycosyltransferase from W. somnifera, in plant's adaptation to abiotic stress. The WsSGT...

Selection of transgenic WsSGTL1 overexpression lines of A.thaliana. (A) T1 generation transgenics in ½ MS media supplemented with 50 µg ml−1 kanamycin. (B) PCR analysis of 12 positive transgenic lines by Gene Specific Primers in T2 generation (amplicon size 2.1 Kb). (TIF)

Spider plots of chlorophyll fluorescence. Spider plots showing relative changes of mean values of selected fluorescence parameters of maximum photosynthetic efficiency (Fv/Fm), photosynthetic yield Y(II), excitation pressure 1-Y(II), total nonphotochemical quenching (NPQ), regulated heat dissipation Y (NPQ), and unregulated heat dissipation Y (NO)...

List of primers used in present study. (DOCX)

Colony PCR: Analysis of WsGTL1 by WsSGTL1 (F) and WsSGTL1(R) primers; C1 negative control (lane 1), C2, C3, C4 and C5 positive colony (lane 2–5). (TIF)

Schematic diagram showing T-DNA region of binary vector pBI121 used for transformation. The WsSGTL1 gene was inserted at the XbaI-SacI site in sense orientation. Promoter DECaMV35S (600 bp), WsSGTL1 (2.1 Kb) and the restriction sites were assembled and transformed in A. thaliana. (TIF)

HPLC chromatogram of standard sterol compounds. HPLC chromatogram of standard sterol with retention time Campesterol: 23.477, Stigma sterol: 24.619, Sito sterol: 28.480 (TIF)

Relative expression of WsSGTL1 through real time PCR. Relative expression of WsSGTL1 in transgenic A.thaliana regulated by CaMV35S promoter under salt, heat and cold stress. Plants were treated with 0, 50, 100 or 150 mM NaCl for 24 h. WsSGTL1 expression was not detected in WT plants grown under both with or without salt stress. While it was markedl...

A Genome walking PCR with adaptor primers and gene specific primers. PCR products from non template control (lane 1), nested control (lane 2), AP1+GSP1 (lane 3), AP2+GSP2 (lane 4), AP1+GSP3 (lane 5) and AP2+GSP4 (lane 6) are shown. M;λ HindIII/EcoRI double digested DNA molecular weight markers. B. Promoter sequence of WsSGTL1. Nucleotide sequence o...

Secondary structure and alignments (A) Secondary structure of WsSGTL1. (B) Secondary structure of AtSGT.(C) Alignment of WsSGTL1 and AtSGT. (ZIP)

Germination rate and early seedling development of both WT and overexpression lines of A.thaliana under salt stress. (A) Percentage of germinating seeds of WT and WsSGTL1 overexpression lines (L1, L2 and L3) of T3 plants grown on MS medium. (B) Supplemented with 50 mM NaCl. (C) With 100 mM NaCl. (D) With 150 mM NaCl. Data of germination recorded af...

Phenotypic differences between WsSGTL1 overexpression lines of A. thaliana and WT (Col 0). (A) Seedlings after 14 days of germination grown with 0 mM NaCl. (B) Seedlings grown after 7 days of germination (from L to R); with 50 mM NaCl; with 100 mM NaCl; with 150 mM NaCl. (TIF)

Effects of different abiotic stress on both genotypes of A. thaliana (WT and WsSGTL1 transgenic lines) in reference to fluorescence parameters. Maximum quantum yield of PSII photochemistry (Fv/Fm), effective quantum yield of PSII Y (II)), total heat dissipation (NPQ), yield of regulated heat dissipation Y (NPQ), and yield of unregulated heat dissip...

Data of phenotypic comparison of WT plants and WsSGTL1 overexpression lines of A. thaliana after 14 days of germination. Comparison of shoot length, root length, shoot weight and leaf count of WT and WsSGTL1 overexpression lines of A. thaliana grown on MS medium supplemented with 50 mM NaCl, 100 mM NaCl and 150 mM NaCl. (A) Average shoot length.(B)...

Preliminary properties of the WsSGTL1 protein predicted by ProtParam. (TIF)

Photosynthetic yield [Y (II)] and total heat dissipation (NPQ) under different abiotic stress conditions. (A) In WT and transgenic lines of A.thaliana, photosynthetic yield decreased at 50 and 100 mM NaCl but in WT its values decreased significantly due to stress (P≤0.01). (B) Total heat dissipation in WT decreased significantly (P≤0.01) but not un...

The aim of the present research work was to develop a protocol for slow growth conservation. Spectacular success was achieved in preserving shoot apices of Glycyrrhiza glabra under slow growth conditions. Cultures responded best, when incubated at 10ºC under low light intensity (2.5 µmol m -2 s -1 quantum flux density) and sealed with polypropylene...

The aim of the present research work was to develop a protocol for slow growth conservation. Spectacular success was achieved in preserving shoot apices of Glycyrrhiza glabra under slow growth conditions. Cultures responded best, when incubated at 10ºC under low light intensity (2.5 µmol m-2 s-1 quantum flux density) and sealed with polypropylene c...

Opium poppy (Papaver somniferum) remains the most important source for several pharmaceutical benzylisoquinoline alkaloids including the narcotic analgesic morphine, the anti-tussive drug codeine. Selection, breeding as well as other molecular practices have produced large number of germplasm with modulated biosynthesis and altered accumulation of...

An improved protocol has been developed for micropropagation of Jatropha curcas to obtain quality planting material on large scale. To obtain improved shoot growth, rooting and acclimatization of in vitro plants, multiplication medium was modified. Shoots multiplied in Murashige and Skoog's (MS) medium containing 15 mg dm−3 each of l-glutamine and...

In the present study, Agrobacterium tumefaciens-mediated transformation protocol has been optimized for Jatropha curcas—an important biofuel plant using leaf segments (LS) and hypocotyl segments (HS) from in vitro-seedlings. Among different accessions used, ‘GJ-Ran-C2’ was selected as the best for transformation. A. tumefaciens strain EHA101, conta...

Sterol glycosyltransferases (SGTs) catalyze the transfer of sugar molecules to diverse sterol molecules, leading to a change in their participation in cellular metabolism. Withania somnifera is a medicinal plant rich in sterols, sterol glycosides and steroidal lactones. Sterols and their modified counterparts are medicinally important and play a ro...

Sterols are important components of cell membranes, hormones, signalling molecules and defense-related biotic and abiotic chemicals. Sterol glycosyltransferases (SGTs) are enzymes involved in sterol modifications and play an important role in metabolic plasticity during adaptive responses. The enzymes are classified as a subset of family 1 glycosyl...

A high yielding elite plant of Jatropha curcas was established under aseptic condition from field-grown nodal explants. Shoots were proliferated in MS medium supplemented with 0.5 mg dm−3 benzyladenine and 0.1 mg dm−3 indolebutyric acid along with 10 mg dm−3 adenine sulphate and a combination of 15 mg dm−3 each of l-glutamine and l-arginine. Howeve...

A variant plant of opium poppy (Papaver somniferum L.) having high thebaine was obtained from the dose of 10kR (kilo rad)+0.4% EMS during an extensive mutation breeding experiment. The M2 seeds of variant plant were subjected to in vitro studies to investigate the prospects of thebaine production through tissue culture. Alkaloid profile of variant...

Gerbera is the most popular ornamental plant of temperate climate but few cultivars grow in subtropical climate. For popularization of this crop in north Indian plains, like Lucknow, tissue culture is the only alternative method to begin with. A lot of work on tissue culture of gerbera has been done but the selection of suitable cultivars growing i...

Leaf explants of the second or third node were collected from field-grown elite Jatropha curcas trees and incubated in Murashige and Skoog’s (Physiol Plant 15:473–497, 1962) medium supplemented with growth regulators. Direct shoot organogenesis was induced when explants were incubated in a medium containing 0.5mgl−1 benzyladenine (BA) and 0.1mgl−1...

A protocol for indirect differentiation of shoots / roots from leaf callus of Asiatic hybrid lily was developed through in vitro methods. The involvement of antioxidant enzymes, like, SOD, POX and CAT, and their isoenzymes during organogenesis in the morphogenetic callus was stud ied.The activity of these enzymes was increased during early developm...

This report describes Agrobacterium tumefaciens-mediated transformation of Withania somnifera--an important Indian medicinal plant. A. tumefaciens strain LBA4404, containing the binary vector pIG121Hm was used for transformation, along with the gusA reporter gene with intron under the transcriptional control of the Cauliflower Mosaic Virus (CaMV) 3...

A protocol for clonal propagation of Rosa clinophylla, a rare and endangered species but very important for breeding purposes had been standardized through in vitro axillary bud culture. Although cytokinins alone were able to induce shoot buds, but their proper growth and number could be increased only when they were used in combination with GA3. H...

A protocol for direct differentiation of shoots from leaf segments of Litium cv ‘Orange Pixie’ was developed through in vitro methods. After hardening, tissue-raised plants were transferred in the open field conditions from the very beginning. The acclimatized plants not only grew well but flowered also at 43°C under subtropical climatic conditions...

'Madam E Roger', a greenish white large flowered chrysanthemum cultivar- was selected for in vitro propagation and mutagenesis to induce further genetic variability. A protocol has been standardized to develop large-scale quality planting material for commercial exploitation. Shoot bud differentiation could be achieved in ray florets in the presenc...

Ray florets of Chrysanthemum morifolium Ramat. cvs. Flirt, Puja, Maghi and Sunil were treated with 500 and 1000 rad gamma rays and cultured on MS medium supplemented with different concentrations and combinations of growth regulators. The frequency of direct shoot regeneration decreased in gamma-ray-treated florets. Radiation effect was found on pl...