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Showing papers on "Kinetin published in 2020"


Journal ArticleDOI
TL;DR: An efficient and reproducible in vitro regeneration protocol was established for chickpea and the in vitro regenerated plants from all four explants were found to be the true to type with their mother plant.
Abstract: High efficient and repeatable in vitro regeneration protocol was established from embryo axis, half-seed, axillary meristem, and cotyledonary node explants of chickpea. Various concentrations and combinations of various plant growth regulators (PGRs) were employed to induce multiple shoots, shoot elongation and rooting of shoots to obtain complete plantlets of chickpea. The pretreatment of seeds with 6-benzyl aminopurine (BAP) at 1.0 mg l−1 was found to significantly increase the multiple shoot regeneration from the all explants tested. Among three PGRs such as BAP, kinetin (KIN) and thidiazuron (TDZ) tested for multiple shoot induction; BAP at 2.0 mg l−1 produced the maximum number of shoots in all tested explants. The maximum number of shoots (48.80 shoots/explant) was attained from the embryo axis explant followed by half-seed (32.76 shoots/explant), axillary meristem (28.34 shoots/explant) and cotyledonary node explant (18.47 shoots/explant) on medium augmented with 2.0 mg l−1 BAP along with 0.05 mg l−1 Indole-3-butyric acid (IBA). The optimum percentage of shoot elongation response was recorded (96.68%) on medium fortified with IAA (0.05 mg l−1), GA3 (1.0 mg l−1) and BAP (1.0 mg l−1) with an average shoot length of 8.82 cm. The elongated shoots were successfully rooted in medium augmented with 2.0 mg l−1 IBA. The complete plants were acclimatized in the greenhouse with a survival rate of 72%. The plantlets regenerated from four explants appeared to be morphologically similar to mother plants. The genetic fidelity of in vitro regenerated plants was evaluated using Start Codon Targeted and Inter simple sequence repeats molecular markers. The in vitro regenerated plants from all four explants were found to be the true to type with their mother plant. The in vitro protocol presented in the study should offer as a feasible system for chickpea genetic transformation. An efficient and reproducible in vitro regeneration protocol was established for chickpea. Application of different concentrations and combinations of PGRs was found to enhance multiple shoot induction, shoot elongation, rooting and acclimatization of in vitro regenerated plants in field conditions, and further evaluated genetic fidelity using molecular markers.

37 citations


Journal ArticleDOI
TL;DR: F foliar micro-morphological characters like stomata, trichome and vein pattern of leaves were studied from in vitro and acclimatized plants to focus on the developmental adaptation of micropropagated plantlets towards survival in field conditions.

33 citations


Journal ArticleDOI
TL;DR: It is concluded that floral buds, leaves and stem could also be used as an alternative potential source for steroidal alkaloids and callus culture as sustainable approach for metabolite production to meet industrial standards and demands.

28 citations


Journal ArticleDOI
TL;DR: The protocol described here is a rapid and reliable micropropagation protocol for large-scale production of clonally stable clary sage plants for commercial purposes.

26 citations


Journal ArticleDOI
TL;DR: The monomorphic pattern of inter simple sequence repeats (ISSR) and start codon targeted (SCoT) markers of in vitro raised plants matched with mother plants confirmed the genetic stability and this method could be efficiently utilized for commercial-scale production of B. balcooa.

23 citations


Journal ArticleDOI
TL;DR: Experimental results on the evaluation of physiological, biochemical parameters showed the role of pigment molecules and antioxidant systems in the production of albino micro shoots and the photoperiodic incubation duration showed 12 h as the best light period and sub or supra-optimal resulted in theproduction of abnormal and albinos micro shoots.
Abstract: Chonemorpha fragrans is an endangered medicinal woody climber, regarded among alternative plant sources of camptothecin Camptothecin is a monoterpene indole anti-cancer alkaloid with annual trade value of over three billion US dollars in the recent, and is used in the production of its analog drugs approved for the chemotherapy of cancer of varied types Effects of plant growth regulators, culture media strength and photoperiodic duration on the micropropagation efficiency of C fragrans from nodal segment explants were studied on Murashige and Skoog (MS) medium amended with Thidiazuron (TDZ), Benzylaminopurine (BAP) or Kinetin (Kin) Thidiazuron was more efficient over BAP and Kin when half basal MS medium was used over full or quarter strength Results of carbon source experiment showed sucrose as the most effective over glucose, fructose, and maltose in the clonal production Studies on the photoperiodic incubation duration showed 12 h as the best light period and sub or supra-optimal resulted in the production of abnormal and albino micro shoots Experimental results on the evaluation of physiological, biochemical parameters showed the role of pigment molecules and antioxidant systems in the production of albino micro shoots

21 citations


Journal ArticleDOI
TL;DR: The aim of this study was to analyze the influence of plant growth regulators (PGRs) on the development, quality, and physiological state of in vitro-grown bleeding heart “Gold Heart” and “White Gold”.
Abstract: There is little information on the in vitro tissue culture systems in Lamprocapnos spectabilis (bleeding heart). The aim of this study was to analyze the influence of plant growth regulators (PGRs) on the development, quality, and physiological state of in vitro-grown bleeding heart “Gold Heart” and “White Gold”. Single-node explants were inoculated on the modified MS medium (Murashige and Skoog in Physiol Plant 15:473–497, 1962), fortified with different auxins, which included indole-3-acetic acid (IAA), 1-naphthaleneacetic acid (NAA), and picloram (PIC), along with cytokinins, which included 6-benzyladenine (BA), kinetin (KIN), and thidiazuron (TDZ) at various concentrations. The morphogenetic response of the explants was cultivar-specific. KIN was preferable for the proliferation and development of shoots in “Gold Heart.” However, none of the auxins or cytokinins improved the development of “White Gold” explants, compared with the PGR-free control medium. NAA was the most effective for stimulating rhizogenesis in both cultivars, although IAA resulted in the regeneration of the longest roots. TDZ, NAA, and PIC suppressed the development of shoots in both cultivars tested and stimulated abundant callus formation. Indirect regeneration of somatic embryos occurred on the NAA- and PIC-fortified media. In particular, the latter media stimulated regeneration of the highest number of somatic embryos per nodal segment. Composition of the culture medium also affected the levels of primary and secondary metabolites in shoots and callus of L. spectabilis. IAA (at 1.0 mg L−1) stimulated the synthesis of chlorophyll a and carotenoids in the “Gold Heart,” while BA and KIN (at 0.5 mg L−1) had a negative impact on the concentration of chlorophyll b in the shoots of this cultivar. None of the PGRs increased the level of the pigments in the shoots of bleeding heart “White Gold.” The concentration of chlorophylls and carotenoids in the callus of both cultivars tested was significantly lower compared with the shoots; however, callus was abundant in flavanols.

21 citations


Journal ArticleDOI
21 Feb 2020
TL;DR: The results indicated that the exogenous application of kinetin increased the total radical scavenging capacity of coffee plants, which has the potential to reinforce antioxidant capacity, as well as modulate the decline in photosynthetic productivity resulting in improved tolerance under cold stress conditions.
Abstract: Coffee plants are seasonally exposed to low chilling temperatures in many coffee-producing regions. In this study, we investigated the ameliorative effects of kinetin-a cytokinin elicitor compound on the nonenzymatic antioxidants and the photosynthetic physiology of young coffee plants subjected to cold stress conditions. Although net CO2 assimilation rates were not significantly affected amongst the treatments, the subjection of coffee plants to cold stress conditions caused low gas exchanges and photosynthetic efficiency, which was accompanied by membrane disintegration and the breakdown of chlorophyll pigments. Kinetin treatment, on the other hand, maintained a higher intercellular-to-ambient CO2 concentration ratio with concomitant improvement in stomatal conductance and mesophyll efficiency. Moreover, the leaves of kinetin-treated plants maintained slightly higher photochemical quenching (qP) and open photosystem II centers (qL), which was accompanied by higher electron transfer rates (ETRs) compared to their non-treated counterparts under cold stress conditions. The exogenous foliar application of kinetin also stimulated the metabolism of caffeine, trigonelline, 5-caffeoylquinic acid, mangiferin, anthocyanins and total phenolic content. The contents of these nonenzymatic antioxidants were highest under cold stress conditions in kinetin-treated plants than during optimal conditions. Our results further indicated that the exogenous application of kinetin increased the total radical scavenging capacity of coffee plants. Therefore, the exogenous application of kinetin has the potential to reinforce antioxidant capacity, as well as modulate the decline in photosynthetic productivity resulting in improved tolerance under cold stress conditions.

19 citations


Journal ArticleDOI
TL;DR: A combination of phytohormones (indole acetic acid and kinetin) was augmented in nitrogen-limited medium to achieve higher biomass and lipid yield in Graesiella emersoniiNC-M1 and Chlorophyta sp.

19 citations


Journal ArticleDOI
Arda Acemi1
TL;DR: It is suggested that well-characterized chitosan could be used as an alternative to JAS and BAP in orchid cultures and induce similar effects with jasmonic acid and 6-benzylaminopurine inOrchid cultures.
Abstract: This study aimed to compare the in vitro effects of chitosan oligomers and polymer with commonly-used plant growth regulators (PGRs) on seed germination, protocorm formation, and organ development in Serapias vomeracea. The effects of N-acetylated (10%) chitosan oligomer mixture (CHI-OM) with a degree of polymerization (DP) between 2 and 15 (5, 10, 15, and 20 mg L−1) and chitosan polymer (CHI-P) with a DP of 70 were compared with commonly-used cytokinins [6-benzylaminopurine (BAP) and kinetin (KIN)], auxins [indole-3-acetic acid (IAA) and indole-3-butyric acid (IBA)], and jasmonic acid (JAS) at 0.25, 0.5, 1.0, and 2.0 mg L−1. The medium supplemented with CHI-P at 5 mg L−1 gave the highest seed germination rate, whereas JAS and CHI-OM triggered protocorm formation better than the all treatments tested. The JAS and IAA treatments resulted in intense browning of the roots. The CHI-P treatments at high concentrations and JAS treatments at moderate concentrations increased mean shoot length while the medium containing KIN at 0.5 mg L−1 induced root elongation significantly. The root elongation-inhibitory effect of chitosan was observed at the higher concentrations of CHI-OM, but the media containing 10 mg L−1 CHI-OM and 15 mg L−1 CHI-P triggered adventitious rooting. However, the highest tuberization success was found after 10 mg L−1 CHI-OM and 0.5 mg L−1 JAS treatments. Tuber development was achieved in the media supplemented with BAP and CHI-OM. This study suggested that well-characterized chitosan could be used as an alternative to JAS and BAP in orchid cultures. Well-characterized chitosans at the right concentration induce similar effects with jasmonic acid and 6-benzylaminopurine in orchid cultures.

19 citations


Journal ArticleDOI
TL;DR: The highest callus induction frequency was observed on MS (Murashige and Skoog, 1962) medium supplemented with 4 µM 6-benzyladenine (BA) after four weeks of culture period, and the efficiency of stable transformation was found to be approximately 7% in the transgenic plants.

Journal ArticleDOI
01 Jun 2020
TL;DR: Phytochemical screenings revealed that the micropropagated tissues exhibited higher contents of secondary metabolites, antioxidant potentials and chlorophyll contents as compared to the mother plant, and genetic stability of the acclimatized plantlets was evaluated.
Abstract: Coelogyne ovalis Lindl. is an evergreen epiphytic, sympodial, ornamental orchid having medicinal properties due to the presence of bioactive compounds such as flavidin, flavidinin, coelogin. In the present study, nodal buds of Coelogyne ovalis, when cultured in Knudson C medium supplemented with meta-Topolin (mT), 6-benzyl aminopurine (BAP), kinetin (Kn) and α-naphthalene acetic acid (NAA), responded to the induction of protocorm-like bodies (PLBs) or shoot buds. The highest numbers of PLBs (22.73 ± 0.47) and shoot (14.53 ± 0.27) per explant were augmented in the medium containing a combination of mT (10 µM) and NAA (0.5 µM). Meta-Topolin in the medium was found to be superior in enhancing the response of explants as compared to BAP and Kn. The best rooting of the shoots was observed in the medium supplemented with 10 µM indole-3-acetic acid. Well-developed plantlets obtained after 20 weeks of culture were acclimatized and transferred to the net house. Genetic stability of the acclimatized plantlets was evaluated and contrasted with the mother plant using two molecular markers, viz. start codon targeted and inter-simple sequence repeats, wherein 5.15% clonal variability was observed. Phytochemical screenings revealed that the micropropagated tissues exhibited higher contents of secondary metabolites, antioxidant potentials and chlorophyll contents as compared to the mother plant. The present study could be of great significance for the conservation and commercial utilization of C. ovalis, an orchid facing threat due to its overexploitation from nature.

Journal ArticleDOI
TL;DR: Results provided an effective method for the regulation of flavonoid biosynthesis in M. chamomilla cell suspension culture, and the use of SMF as a tool for the induction of apigenin production.
Abstract: This study represents an optimized protocol for callus establishment and cell suspension culture of Matricaria chamomilla, and the impact of the static magnetic field (SMF) on flavonoid metabolism and antioxidant activity were examined for the first time. The effect of growth regulators was investigated to enhance biomass growth and apigenin production. Murashige and Skoog medium supplemented with 2,4-D (1.5 mg l−1) and Kinetin (0.5 mg l−1) showed the highest callus induction rate (100%), fresh weight, apigenin (0.82%) and apigenin-7-glucoside (1.57%) contents. Cell suspension culture was established, and the optimum subculture time was found to 13–15 days. SMF induced cell leaching and oxidative stress in all treated cells by an increase in H2O2 content and more stimulation of superoxide dismutase (SOD), catalase (CAT) and peroxidase (POX) enzymes activities. Total phenolic, flavonoid and DPPH activity increased in cells treated to SMF, and the maximum content of apigenin (1.3%) and apigenin-7-glucoside (2.1%) were identified in cell treated to 4 mT. These results provided an effective method for the regulation of flavonoid biosynthesis in M. chamomilla cell suspension culture, and the use of SMF as a tool for the induction of apigenin production. Cell suspension cultures of Matricaria chamomilla contain valuable medicinal flavonoids. Static magnetic field promoted apigenin production and antioxidative enzyme activities in M. chamomilla cell suspension.

Journal ArticleDOI
TL;DR: This analysis suggests that kinetin concentrations of 500 nM and above cause cytotoxicity as well as genotoxicity in various cell types, but concentrations below 100 nM do not cause any toxicity, and deciphers vital residues in adenine phosphoribosyltransferase and adenosine receptor that facilitate the binding of Kinetin to these two important human cellular proteins.
Abstract: Metabolism and signaling of cytokinins was first established in plants, followed by cytokinin discoveries in all kingdoms of life. However, understanding of their role in mammalian cells is still scarce. Kinetin is a cytokinin that mitigates the effects of oxidative stress in mammalian cells. The effective concentrations of exogenously applied kinetin in invoking various cellular responses are not well standardized. Likewise, the metabolism of kinetin and its cellular targets within the mammalian cells are still not well studied. Applying vitality tests as well as comet assays under normal and hyper-oxidative states, our analysis suggests that kinetin concentrations of 500 nM and above cause cytotoxicity as well as genotoxicity in various cell types. However, concentrations below 100 nM do not cause any toxicity, rather in this range kinetin counteracts oxidative burst and cytotoxicity. We focus here on these effects. To get insights into the cellular targets of kinetin mediating these pro-survival functions and protective effects we applied structural and computational approaches on two previously testified targets for these effects. Our analysis deciphers vital residues in adenine phosphoribosyltransferase (APRT) and adenosine receptor (A2A-R) that facilitate the binding of kinetin to these two important human cellular proteins. We finally discuss how the therapeutic potential of kinetin against oxidative stress helps in various pathophysiological conditions.

Journal ArticleDOI
01 Jan 2020
TL;DR: The present micropropagation protocol could be effectively employed to generate true to type plantlets of C. zedoaria and genetic homogeneity of in vitro plants was further confirmed through ISSR and flow cytometry analysis.
Abstract: The present investigation was carried out to establish an efficient and reproducible micropropagation protocol for the production of morphologically, genetically and chemically uniform plants of Curcuma zedoaria. Axillary bud explants of C. zedoaria were inoculated into MS basal medium supplemented with various combinations and concentrations of 6-benzyladenine (2.2–22.2 µM, BA), kinetin (2.3–23.2 µM, Kin), indole-3-acetic acid (2.9–11.4 µM, IAA), α-naphthalene acetic acid (2.7–10.2 µM, NAA) and adenine sulphate (33.9–203.6 µM, Ads). Almost 95% of rhizome buds sprouted on MS medium supplemented with 13.3 μM BA, 5.7 μM IAA and 63.9 μM Ads giving rise to an average of 12.89 ± 0.02 shoots within 6 weeks. However, the maximum number of roots (25.8 ± 0.07 roots per explant) was obtained on half strength MS medium supplemented with 7.4 µM of IBA after 4 weeks of inoculation. Morphological characteristics were similar in both conventionally propagated and micropropagated plants. Additionally, genetic homogeneity of in vitro plants was further confirmed through ISSR and flow cytometry analysis. A total of 27 ISSR primers were screened, out of which 13 ISSR primers generated 58 monomorphic and reproducible bands thereby confirming the genetic uniformity of obtained plants. The mean 2C DNA content of the mother plant (2.96 pg) was similar to that of in vitro derived plants (3.07 pg). Gas chromatography-mass spectrometry (GC–MS) analysis showed similarity in the qualitative profile of chemical constituents of essential oil and high-performance liquid chromatography analysis revealed no significant differences in curcumin content in the tissue culture regenerants and mother plants of C. zedoaria. Therefore, the present micropropagation protocol could be effectively employed to generate true to type plantlets of C. zedoaria.

Journal ArticleDOI
TL;DR: The main objective of this study was to develop in vitro systems utilizing N. damascena seedlings, as an easily accessible explant source, for efficient callus induction and proliferation, and plant regeneration via somatic embryogenesis, and to validate the usefulness of the obtained callus as a source of protoplasts and their capability to develop into plants.
Abstract: In this study we report the development of effective in vitro systems for a medicinal plant Nigella damascena L. comprising: (1) callus induction, (2) somatic embryogenesis in callus cultures with subsequent plant regeneration, and (3) isolation and regeneration of callus-derived protoplasts. Callus development was achieved on 83–100% of hypocotyl and cotyledon explants, whereby Murashige and Skoog medium (MS) supplemented with 3 mg L−1 6-benzylaminopurine and 0.5 mg L−1 α-naphthaleneacetic acid (NAA; BN medium) was more advantageous than MS with kinetin and NAA (KN medium). Histological observations of calli revealed the presence of embryogenic zones from which somatic embryos developed on the hormone-free medium. Plant regeneration was observed on 76–95% of calli. A high capacity to form somatic embryos and regeneration was maintained in long-lasting cultures, i.e. even in 2 year old callus. The obtained callus was also a good source tissue for protoplast isolation. By applying a mixture of cellulase and pectolyase, the acceptable yield of viable protoplasts was achieved, especially from hypocotyl-derived callus maintained on BN medium. Protoplasts embedded in an alginate matrix and cultured in modified Kao and Michayluk media re-constructed their cell wall and re-entered mitotic divisions. About 30% of small cell aggregates formed microcalli, which, after the release from alginate, proliferated continuously on KN and BN media, irrespective of the tissue variant used as the protoplast source. Somatic embryo formation and plant regeneration were successful on hormone-free media. An effective plant regeneration system of N. damascena protoplast cultures has been developed and is being reported for the first time. The main objective of this study was to develop in vitro systems utilizing N. damascena seedlings, as an easily accessible explant source, for efficient callus induction and proliferation, and plant regeneration via somatic embryogenesis. Moreover, we attempted to validate the usefulness of the obtained callus as a source of protoplasts and their capability to develop into plants.

Journal ArticleDOI
TL;DR: The direct and indirect somatic embryogenesis protocols reported here having importance in genetic improvement of a cardio-tonic herb D. lanata using recombinant DNA technology is confirmed.

Journal ArticleDOI
01 Mar 2020
TL;DR: In this article, an efficient regeneration protocol for Tecoma stans L, a valuable antidiabetic plant, using shoot tip explants was developed, where different concentrations of benzyl adenine (BA), kinetin (Kin) or isopentyl acetic acid (2-iP) were evaluated for in vitro shoot bud induction and proliferation.
Abstract: The present investigation was carried out to develop an efficient regeneration protocol for Tecoma stans L., a valuable antidiabetic plant, using shoot tip explants. The effect of different concentrations (1.0–10.0 µM) of benzyl adenine (BA), kinetin (Kin) or isopentyl adenine (2-iP) either alone or in combination with different concentrations (0.1–2.0 µM) of α-naphthalene acetic acid (NAA) was evaluated for in vitro shoot bud induction and proliferation. Of the tested concentrations, MS medium containing 7.5 µM BA + 0.5 µM NAA proved to be optimal for the maximum (91%) regeneration with mean shoot number (9.8 ± 0.48) and length (4.36 ± 0.33 cm) after 8 weeks of incubation. The best in vitro rooting in regenerated microshoots was achieved on MS medium supplemented with 1.0 µM IBA, which produced a mean root number (6.8 ± 0.58) and length (4.24 ± 0.29 cm) after 4 weeks of incubation. The regenerated plantlets with well-developed roots and shoots were successfully acclimatized in thermocol cups filled with soilrite, for 8 weeks, in growth chamber. Thereafter transfered to field conditions where they grew well in garden soil with 80% survival rate. During the acclimatization period (0–56 days), a subsequent increase in the content of photosynthetic pigments (chlorophyll a, b and carotenoid) was observed up to 56th day. On assessing genetic fidelity among regenerated plants through RAPD analysis, no polymorphism in banding pattern was observed with that of donor plant, thus indicating clonal stability among micropropagated plants.

Journal ArticleDOI
TL;DR: Histological analysis confirmed the bipolar organization of the somatic embryos, which were comprised of shoot and root meristems, and showed that SE was characterized by globular, heart-shaped, torpedo-shaped and cotyledon stages in O. henryi, the developmental process of which was similar to that of zygotic embryos.
Abstract: An efficient in vitro plant regeneration method via somatic embryogenesis has been established in Ormosia henryi Prain, which is an important precious tree in the timber industry. Five types of explants, six types of basic media and different plant growth regulators (PGRs) were used for embryogenic callus (EC) initiation, EC proliferation, somatic embryo (SE) induction and SE germination. The EC and SE induction rates from mature zygotic embryos (MZE) were higher than those from stem segments, cotyledons, leaves and hypocotyls. MZE were used as explants, and the EC induction rate was greater than 30% on B5 medium supplemented with 1.0 mg/l 6-benzylaminopurine (BA) and 0.5 mg/l naphthaleneacetic acid (NAA) or 0.2 mg/l BA and 2.0 mg/l 2,4-dichlorophenoxyacetic acid (2,4-D). The highest induction rate (22.83%) for SE was obtained with B5 medium supplemented with 0.5 mg/l kinetin (KT) and 1.0 mg/l 2,4-D. The EC proliferation rate and SE number in the suspension culture were increased compared to those in the solid culture. Fifty-four percent of SE developed to the cotyledonary stage on B5 medium supplemented with 0.5 mg/l BA and 0.2 mg/l NAA. The number of cotyledon embryos was the highest on medium supplemented with 0.5 mg/l thidiazuron (TDZ) and 0.2 mg/l NAA. Histological analysis confirmed the bipolar organization of the somatic embryos, which were comprised of shoot and root meristems, and showed that SE was characterized by globular, heart-shaped, torpedo-shaped and cotyledon stages in O. henryi, the developmental process of which was similar to that of zygotic embryos. An efficient in vitro plant regeneration method via somatic embryogenesis has been established in Ormosia henryi Prain using mature zygotic embryos as explant.

Journal ArticleDOI
TL;DR: Although efficient plant regeneration was achieved in the presence of all four tested cytokinins in the three organogenetic pathways, distinct differences exist in terms of shoot regeneration potential.
Abstract: The present study describes the first successful report on an efficient in vitro protocol for direct and indirect organogenesis of an important medicinal and anticancerous plant, Solanum erianthum D. Don. An optimal response using leaf explants was achieved in half-strength Murashige and Skoog (MS)-medium supplemented with 5 mg L−1 naphthalene acetic acid for friable callus formation and 2 mg L−1 naphthaleneacetic acid and 0.5 mg L−1 6-benzylaminopurine amino purine for induction of compact callus. The maximum number of shoot buds (20.33 ± 11.55) was obtained from compact callus using indirect shoot organogenesis in MS medium supplemented with 0.5 mg L−1 thidiazuron among the four cytokinins tested, which included 6-benzyl amino purine, kinetin, 6-γ,γ-dimethylallylamino purine, and thidiazuron. Supplements of 3 mg L−1 6-benzyl amino purine played a crucial role in direct shoot bud formation from a leaf. The buds appeared as small protuberances, and finally developed into leafy shoots as the culture period progressed. Regenerated shoot buds were transferred to MS media that contained four cytokinins for shoot tip culture, in which 8 mg L−1 6-γ,γ-dimethylallylamino purine exhibited the best response (15 ± 2.52). Although efficient plant regeneration was achieved in the presence of all four tested cytokinins in the three organogenetic pathways, distinct differences exist in terms of shoot regeneration potential. Half-strength MS basal medium is recommended for root induction. An approximate 70% survival rate of regenerated plantlets was recorded following sequential transfer from culture conditions to the field. The genetic fidelity of the regenerated plants was evaluated using random amplified polymorphic DNA molecular markers, which showed a high number of uniform monomorphic bands.

Journal ArticleDOI
TL;DR: It is demonstrated that supplementation of buffer 2 with 50 μM kinetin is ideal for reducing the magnitude of oxidative damage during semen cryoprocessing and improving the post-thaw quality of dog semen.

Journal ArticleDOI
TL;DR: Simple adoption, higher culture regeneration and simultaneous production of rooted plantlets in a cyclic manner render the protocol useful for mass scale propagation of elite genotype of female date palm.
Abstract: Phoneix dactylifera L. commonly called date palm is a highly valuable horticultural cash crop for arid and semi-arid regions. The availability of offshoots and their survival during plantation are major concern. Being dioecious tree, seed propagation in date palm do not produce true-to-type offspring and tissue culture propagation is the only viable option to supply quality-planting propagules. Hereby, we report callus culture and plantlet regeneration in female date palm using in vitro-derived adventitious shoot bud tissues as explants. Explants (89.33 ± 2.67%) produced callus culture on 0.8% agar-gelled Murashige and Skoog’s basal medium containing 100.0 mg l−1 each polyvinylpyrrolidone, ascorbic acid and glutamine, 50.0 mg l−1 each citric acid, adenine sulphate and l-arginine as additives, 0.1% activated charcoal (AC), 100 mg l−1 2,4-dichlorophenoxyacetic acid (2,4-D) and 3.0 mg l−1 2-isopentenyladenine (2-iP). Callus culture were amplified on medium containing 3.0 mg l−1 2-iP along with 50 mg l−1 2,4-D for 2 passages and 10 mg l−1 2,4-D for 2 passages. Cultures grew moderately, organized and subsequently regenerated into shoot bud like structures during gradual transfer from medium containing higher concentration of 2,4-D to lower concentration. Plantlets were developed by sub-culturing of differentiated buds on (1) hormone free medium supplied with 10.0% sucrose and (2) medium containing 100.0 mg l−1 each ascorbic acid and glutamine, 50.0 mg l−1 each citric acid, adenine sulphate and l-arginine as additives, 1.0 mg l−1 each 6-benzylaminopurine, kinetin, 2-iP and α-naphthaleneacetic acid. Plantlets were developed on medium containing 0.1% AC, 1.0 mg l−1 each indole-3-acetic acid and indole-3-butyric acid. Rooted plantlets were soil-transplanted and acclimatized through gradual exposure from in vitro to in vivo conditions. Simple adoption, higher culture regeneration and simultaneous production of rooted plantlets in a cyclic manner render the protocol useful for mass scale propagation of elite genotype of female date palm.

Journal ArticleDOI
TL;DR: It was found that B5 medium with 1 mg/L 2,4‐D along with 2 g/L peptone produced more leaf callus biomass and enhanced production of podophyllotoxin, kaempferol, and quercetin compared to control and D. pleiantha callogenesis can provide an alternative source forEnhanced production of secondary compounds regardless of the exploitation of its natural plant population.
Abstract: Dysosma pleiantha (Hance) Woodson is one of the endangered traditional Chinese medicinal herbs, highly valued for its medicinal properties by Taiwan's mountain tribes. The present study aims to develop an efficient protocol for callus biomass by optimizing suitable culture medium, carbon source culture condition, and enhanced production of pharmaceutically important podophyllotoxin, kaempferol, and quercetin from callus culture of D. pleiantha under the influence of different additives. Best callus induction was achieved in Gamborg's medium (B5) with 1 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D) along with 0.2 mg/L kinetin under dark condition. Tender leaves of D. pleiantha showed the maximum of 86% callus induction among the different explants tested. Highest leaf callus proliferation was noted in B5 medium with 1 mg/L 2,4-D incubated under complete darkness. In addition, it was found that B5 medium with 1 mg/L 2,4-D along with 2 g/L peptone produced more leaf callus biomass and enhanced production of podophyllotoxin (16.3-fold), kaempferol (12.39-fold), and quercetin (5.03-fold) compared to control. Therefore, D. pleiantha callogenesis can provide an alternative source for enhanced production of secondary compounds regardless of the exploitation of its natural plant population.

Journal ArticleDOI
TL;DR: A significant, improved, and repeatable micropropagation protocol of M. parvifolia using nodal explants of a mature tree is reported and the first report of concurrent ex vitro rooting and acclimatization (CEVRA) in this species is reported.
Abstract: Mitragyna parvifolia (Roxb.) Korth., commonly known as “Kadam,” is an endangered and pharmaceutically valued tree of the family Rubiaceae. The numerous medicinal properties are attributed to the various alkaloids of this plant. Poor seedling survival (due to very small size of seeds, approximately 10,000 per gm), overexploitation and habitat destruction are the major constraints in conserving the wild stocks of this species. This paper reports a significant, improved, and repeatable micropropagation protocol of M. parvifolia using nodal explants of a mature tree. Nodal explants harvested during spring season from the lopped tree differentiated the maximum number of axillary shoots (5.3 ± 0.82 per node) on full-strength Murashige and Skoog (MS) medium containing 3.0 mg L−1 6-benzylaminopurine (BAP) and additives (25 mg L−1 each of adenine sulfate, L-arginine, and citric acid and 50 mg L−1 ascorbic acid). Shoots were amplified in vitro through (1) recurrent transfer of mother explants and (2) subculturing on fresh nutrient medium. The greatest number of shoots (13.4 ± 1.26) with an average length of 6.2 ± 1.03 cm was produced after 4 wk on MS medium containing 0.5 mg L−1 BAP, 0.25 mg L−1 kinetin (Kin), 0.1 mg L−1 Indole-3-acetic acid (IAA), additives, 100 mg L−1 activated charcoal (AC), and 0.8% (w/v) agar. This is the first report of concurrent ex vitro rooting and acclimatization (CEVRA) in M. parvifolia. About 90% micropropagated shoots rooted ex vitro on pulse treatment of 500 mg L−1 Indole-3-butyric acid (IBA; for 5 min) and produced 8.5 ± 0.97 roots per shoot with an average length of 9.40 ± 1.06 cm, after 5 wk. Over 80% of CEVRA plantlets were successfully transplanted to the soil in field. The defined protocol can be employed for conservation ex situ and restoration/rehabilitation/reintroduction in situ of M. parvifolia.

Journal ArticleDOI
TL;DR: The results demonstrated that glutathione-S-transferase (GSTs) was involved in fighting heavy metal stress, and might also have a role in in vitro regeneration of M. pruriens, a fast growing tropical medicinal legume.
Abstract: The effect of heavy metals, CuSO4 and ZnSO4 on morphogenic response of cotyledonary node explants of Mucuna pruriens (L.), a fast growing tropical medicinal legume was evaluated. The climber holds economic importance in traditional as well as modern pharmaceutical areas. Murashige and Skoog (Physiol Plant 15:473–497, 1962) medium supplemented with various strength (0.5, 2.5, 4.5, 6.5 or 8.5 µM) of BA (6-benzyladenine), Kn (Kinetin) and 2-iP (2-isopentenyl adenine) were screened to establish optimum culture conditions for in vitro regeneration using cotyledonary node explant. Optimized medium (MS + BA 2.5 µM) was augmented with different concentration of CuSO4 (0.5–25.0 µM) and ZnSO4 (5–150 µM) to evaluate the effect of heavy metals on regeneration potential of the explants. Among various concentrations tested, MS medium with optimized BA (2.5 µM) along with 5.0 µM CuSO4 or 80.0 µM ZnSO4 proved beneficial, inducing 11.20 and 13.40 shoots per explant, respectively. The results demonstrated that glutathione-S-transferase (GSTs) was involved in fighting heavy metal stress, and might also have a role in in vitro regeneration. Elongated healthy microshoots on transfer to half strength MS medium produced 3.40 roots per shoot after 28 days of incubation without the necessity of exogenous auxin. The in vitro grown regenerants were hardened in Soilrite, and then established in garden and sandy soil (1:1) under natural light where they showed normal growth and development. The work highlights the enhanced multiplication and better growth of the regenerants using elevated concentration of CuSO4 and ZnSO4. GSTs overexpression activity not only confers to overcome the stress but enhanced the in vitro regeneration of M. pruriens also.

Journal ArticleDOI
TL;DR: Visual assessment of micropropagated plants in the field did not show any off-types, based on gross morphological changes of plant morphology or disease reaction, compared to plants of the same genotype derived from a traditional propagation method (stem cuttings).
Abstract: The objective of this study was to set up a plant micropropagation facility to mass propagate sugarcane, energy cane, and related clonally propagated species. An efficient methodology for micropropagation of energy cane and perennial grasses using temporary immersion bioreactors was developed. Several different methods of tissue culture initiation, multiplication, and rooting were evaluated for several varieties of sugarcane (Saccharum officinarum L.) and sugarcane-related species such as Erianthus spp., Miscanthus spp., and Sorghum spp. × sugarcane hybrids, all from a germplasm collection. Apical meristem cultures were initiated for all genotypes that were micropropagated, when liquid or semisolid Murashige and Skoog (MS) medium was used, which was supplemented with 0.1–0.2 mg L−1 BAP, 0.1 mg L−1 kinetin, 0–0.1 mg L−1 NAA, and 0–0.2 μg L−1 giberellic acid. These cultures produced shoots between 4 and 8 wk after initiation. Shoot regeneration from leaf rolls or immature inflorescences was observed as early as 4 wk after initiation. Shoot multiplication was successful for all genotypes cultured in MS medium with 0.2 mg L−1 BAP and 0.1 mg L−1 kinetin. Energy cane had a significantly higher combined multiplication rate when grown under four or five LED lamps than when grown under three LED lamps, or under fluorescent lights in a growth chamber. The addition of 2 mg L−1 NAA produced faster and better rooting in all of the genotypes tested. Shoots produced well-developed roots after one cycle of 15–21 d in the bioreactors. The maximum number of plantlets produced per bioreactor was 1080. Plantlets developed a vigorous root system and were ready to be transplanted into the field after 2 mo. A protocol was standardized for different energy cane clones that were recommended for their biomass production and cell wall composition. Different tissues were used to speed up or facilitate tissue culture initiation. Visual assessment of micropropagated plants in the field did not show any off-types, based on gross morphological changes of plant morphology or disease reaction, compared to plants of the same genotype derived from a traditional propagation method (stem cuttings). This is the first report of energy cane and Miscanthus spp. micropropagation using the SETIS bioreactor.

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TL;DR: Results establish that cypermethrin induces toxicity on photosynthesis, photosynthetic pigments and growth, and this effect was more pronounced in Anabaena PCC 7120 than Nostoc muscorum ATCC 27893.

Journal ArticleDOI
03 Mar 2020
TL;DR: Establishment of an efficient and successful protocol for direct plant regeneration of white lupin from different cotyledonary nodes is described and sucrose showed the highest regeneration frequency with HCN.
Abstract: The tissue culture regeneration system of Lupinus albus has always been considered as recalcitrant material due to its genotype-dependent response and low regeneration efficiency that hamper the use of genetic engineering. Establishment of repeatable plant regeneration protocol is a prerequisite tool for successful application of genetic engineering. This aim of this study was to develop standardized, efficient protocol for successful shoot induction from cotyledonary node of white lupin. In this study, 5 day old aseptically cultured seedlings were used to prepare three explants (half cotyledonary node, HCN; whole cotyledonary node, WCN; and traditional cotyledonary node, TCN), cultured on four concentrations of M519 medium (M519, ½ M519, 1/3 M519, and ¼ M519), containing four carbohydrate sources (sucrose, fructose, maltose, and glucose), and stimulated with various combinations of KT (kinetin), and NAA (naphthalene acetic acid) for direct shoot regeneration. High frequency of 80% shoot regeneration was obtained on ½ M519 medium (KT 4.0 mg L-1 + NAA 0.1 mg L-1) by using HCN as an explant. Interestingly, combinations of (KT 4.0 mg L-1 + NAA 0.1 mg L-1 + BAP 1.67 mg L-1), and (KT 2.0 mg L-1 + NAA 0.1 mg L-1) showed similar shoot regeneration frequency of 60%. Augmentation of 0.25 g L-1 activated charcoal (AC) not only reduced browning effect but also improved shoot elongation. Among the all carbohydrate sources, sucrose showed the highest regeneration frequency with HCN. Additionally, 80% rooting frequency was recorded on ½ M519 containing IAA 1.0 mg L-1 + KT 0.1 mg L-1 (indole acetic acid) after 28 days of culturing. The present study describes establishment of an efficient and successful protocol for direct plant regeneration of white lupin from different cotyledonary nodes.

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TL;DR: Results demonstrate that exogenously supplied PAs improve plant regeneration using somatic embryogenesis and A. tumefaciens-mediated transformation of embryogenic callus of sugarcane ‘Co 86032’, which were more than twofold higher than the control treatment.
Abstract: The influence of exogenous polyamines (PAs) on somatic embryogenesis from immature leaf roll explants and Agrobacterium tumefaciens-mediated transformation of embryogenic callus of Saccharum spp. (sugarcane) ‘Co 86032’ was examined. Immature leaf roll-derived embryogenic callus was obtained on Murashige and Skoog with Gamborg B5 vitamins (MSB5) medium containing 3 mg L−1 2,4-dichlorophenoxyacetic acid (2,4-D). Various concentrations of PAs along with 2 mg L−1 2,4-D and 0.5 mg L−1 kinetin (Kin) were tested for somatic embryo induction. A total of 106 somatic embryos per 250 mg of callus (96.3% responsive explants) were obtained on medium supplemented with 20 mg L−1 putrescine (PUT) and 92.0% of the somatic embryos matured and produced 98 shoots per 250 mg of callus. Somatic embryo induction and maturation was increased more than two- and threefold, respectively, on PUT-supplemented medium compared to control cultures. Histomorphological analyses of various developmental stages verified somatic embryogenesis from immature leaf roll explants. The rooted plantlets were successfully hardened and exhibited normal growth. The efficiency of A. tumefaciens-mediated transformation of embryogenic callus using various concentrations of PAs in the infection, co-cultivation, and regeneration media was also assessed. Putrescine at 20 mg L−1 showed the highest regeneration (54.4%) and transformation (35.8%) efficiencies, which were more than twofold higher than the control treatment. These results demonstrate that exogenously supplied PAs improve plant regeneration using somatic embryogenesis and A. tumefaciens-mediated transformation of embryogenic callus of sugarcane ‘Co 86032’.

Journal ArticleDOI
TL;DR: Findings revealed that synergistic utilization of KN and PUT modulate growth and biochemical processes in seedlings efficaciously in comparison to the individual application under salt stress, and it may be due to a regulatory crosstalk mechanism.
Abstract: Salinity is one of the most vicious environmental constraints that hamper agricultural production. Experiments were done to explore the significant role of sole and synergistic supplementation of kinetin (100 µM KN) and putrescine (100 µM PUT) on Luffa acutangula in NaCl (100 mM) treatment. The harmful effects of salinity on growth were manifested by decreased seedling length, biomass, and pigment contents. We studied the effect of KN, and PUT in preventing salt (NaCl) induced physiological disorders and oxidative damages in 20-day-old Luffa acutangula seedlings. The individual application of KN and PUT increased growth and biochemical parameters, whereas combined KN + PUT treatment showed significant enhancement in growth, photosynthetic pigment content, and osmolyte accumulation in salt-affected plants. Application of KN and PUT also prevented hydrogen peroxide and superoxide production as confirmed by inhibition in electrolyte leakage and lipid peroxidation. Kinetin and PUT application upregulated the antioxidant defense system by enhancing antioxidant enzymes and non-enzymatic contents. Luffa seedlings treated with NaCl + KN + PUT showed 79, 26, 74, and 73% rise in superoxide dismutase, catalase, ascorbate peroxidase, and glutathione reductase enzymes, respectively, in comparison to NaCl-stressed Luffa acutangula. Findings revealed that synergistic utilization of KN and PUT modulate growth and biochemical processes in seedlings efficaciously in comparison to the individual application under salt stress, and it may be due to a regulatory crosstalk mechanism.