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.

Rapid plant regeneration in industrially important Curcuma zedoaria revealing genetic and biochemical fidelity of the regenerants

Results provide significant comparison of leaf anatomy, pigment content, antioxidant response and phenolic profile between individuals from miscellaneous populations and describe unified cultivation protocols for further research on stress biology.
 The plant communities growing on heavy metal-polluted areas have attracted considerable attention due to their unique ability to tolerate enormous amounts of toxic ions. Three ecotypes of Silene vulgaris representing calamine (CAL), serpentine (SER) and non-metallicolous (NM) populations were evaluated to reveal specific adaptation traits to harsh environment. CAL leaves presented a distinct anatomical pattern compared to leaves of SER and NM plants, pointing to their xeromorphic adaptation. These differences were accompanied by divergent accumulation and composition of photosynthetic pigments as well as antioxidant enzyme activity. In CAL ecotype, the mechanism of reactive oxygen species scavenging is based on the joint action of superoxide dismutase and catalase, but in SER ecotype on superoxide dismutase and guaiacol-type peroxidase. On the contrary, the concentration of phenylpropanoids and flavonols in the ecotypes was unchanged, implying the existence of similar pathways of their synthesis/degradation functioning in CAL and SER populations. The tested specimens showed genetic variation (atpA/MspI marker). Based on diversification of S. vulgaris populations, we focused on the elaboration of similar in vitro conditions for synchronous cultivation of various ecotypes. The most balanced shoot culture growth was obtained on MS medium containing 0.1 mg l−1 NAA and 0.25 mg l−1 BA, while the most abundant callogenesis was observed on MS medium enriched with 0.5 mg l−1 NAA and 5.0 mg l−1 BA. For the first time, unified in vitro protocols were described for metallophytes providing the opportunity to conduct basic and applied research on stress biology and tolerance mechanisms under freely controlled conditions.

/pdf/structural-physiological-and-genetic-diversification-of-4md5k6crhv.pdf

Structural, physiological and genetic diversification of Silene vulgaris ecotypes from heavy metal-contaminated areas and their synchronous in vitro cultivation.

Highly sensitive and selective functional nanobiobreaksensors are being developed because they have significant applications in the sustenance and conservation of natural resources and can be used in projects to identify degraded and contaminated areas (of both soil and water) and as environmental quality indicators. In the present study, a nanobiosensor was developed based on using theoretical models (molecular docking and molecular dynamics simulations) based on biomimicry of the action mechanism of herbicides in plants coupled with atomic force microscopy (AFM) tools. The herbicide molecules were detected at very low concentrations using a unique sensor construction: the AFM probes and the substrate were chemically functionalized to favor covalent bonding and promote molecular flexibility, as well as to achieve reproducible and accurate results. Computational methods were used to determine the binding energies associated with the enzyme-herbicide interactions, which were compared with experimental results for adhesion forces. The theoretical results showed that the diclofop herbicide could be assembled and attached onto the mica substrate surface and the ACCase enzyme on the AFM probe without damaging the diclofop molecule. The experimental results showed that using a specific agrochemical target molecule was more efficient than using other nonspecific agrochemicals. On average, there was a 90% difference between the values of specific recognition (diclofop) and nonspecific recognition (imazaquin, metsulfuron, and glyphosate). This result validated the selectivity and specificity of the nanobiosensor. The first evidence of diclofop detection by the AFM probe sensors has been presented in this paper.

https://bv.fapesp.br/pt/publicacao/pdf/91351/nanobiosensor-for-diclofop-detection-based-on-chemically-mod

Nanobiosensor for Diclofop Detection Based on Chemically Modified AFM Probes

Antitumor agents are used in therapy against many forms of human cancer. One of these is mitomycin-C (MMC). As with many agents, it can interact with biological molecules and can induce genetic hazards in non-tumor cells. One of the possible approaches to protect DNA from this damage is to supply antioxidants that can remove free radicals produced by antitumor agents. Lipoic acid (LA) is known as one of the most powerful antioxidants. The aim of this study was to investigate antigenotoxic effects of LA against MMC induced chromosomal aberrations (CA), sister chromatid exchanges (SCE) and micronucleus (MN) formation in human lymphocytes. Lymphocytes were treated with 0.2 μg MMC/heparinized mL for 48 h. Three different concentrations (0.5, 1, 2 μg/mL) of LA were used together with MMC in three different applications; 1 h pre-treatment, simultaneous treatment and 1 h post-treatment. A negative, a positive and a solvent control were also included. In all the cultures treated with MMC + LA, the frequency of abnormal cells and CA/cell significantly decreased compared to MMC. Statistically significant reduction was also observed in SCE/cell and MN frequencies in all treatments. These results demonstrated anticlastogenic and antimutagenic effects of LA against MMC induced genotoxicity. LA showed the most efficient effect during 1 h pretreatment. On the other hand, MMC + LA treatments induced significant reduction in mitotic index than that of MMC treatment alone. These results are encouraging that LA can be a possible chemopreventive agent in tumorigenesis in both cancer patients and in health care persons handling anti-cancer drugs.

/pdf/antigenotoxic-effect-of-lipoic-acid-against-mitomycin-c-in-1jtxfzdaxm.pdf

Antigenotoxic effect of lipoic acid against mitomycin-C in human lymphocyte cultures

Trazodone and milnacipran are the active antidepressant drugs that are being used in the treatment of psychiatric disorders. In this study, the in vitro genotoxic effects of trazodone and milnacipran have been determined in human peripheral blood lymphocytes by using chromosomal aberrations (CAs), sister chromatid exchanges (SCEs), micronuclei (MN), and comet assays. 3.13; 6.25; 12.50; 25.00; 50.00; and 75.00 μg/mL concentrations of trazodone and 2.50; 5.00; 10.00; 20.00; 30.00; and 40.00 μg/mL concentrations of milnacipran were used. Trazodone and milnacipran significantly increased the frequency of CAs and SCEs compared with the control. Both of the active ingredients raised the MN frequency in a dose-dependent manner. Mitotic index was significantly decreased, but replication and nuclear division indices were not affected at all treatments. Trazodone was statistically increased the mean comet tail intensity, tail length, and tail moment at three concentrations (6.25; 12.50; and 25.00 μg/mL) compared with control. Two highest concentrations (50 and 75 μg/mL) of trazodone were toxic in the comet assay. Milnacipran increased the comet tail intensity, tail length, and tail moment at all concentrations. It is concluded that trazodone and milnacipran have clastogenic, mutagenic, and cytotoxic effects on human lymphocytes in vitro.

Evaluation of cytogenetic and DNA damage induced by the antidepressant drug-active ingredients, trazodone and milnacipran, in vitro.

Diclofop-methyl (DM) is a chlorophenoxy derivative used in large quantities for the control of annual grasses in grain and vegetable crops. In this study, the genotoxic effects of DM were investigated by measuring chromosomal aberrations (CAs) in mouse bone-marrow cells and CA and the comet assay in human peripheral lymphocytes. Mice were treated with 15.63, 31.25, 62.5, and 125 mg/kg body weight of DM intraperitoneally for 24 hours, and 15.63-, 31.25-, 62.5-, 125-, and 250-µg/mL concentrations were applied to human lymphocytes for both 24 and 48 hours. In in vivo treatments, DM significantly, but not dose dependently, increased the total chromosome aberrations, compared to both negative and solvent controls. Cell proliferation was significantly, but not dose dependently, affected by all doses. In in vitro treatments, DM (except 15.63 µg/mL) significantly and dose dependently increased the frequency of chromosome aberrations. Also, 250 µg/mL of 48-hour treatment was found to be toxic. Cell proliferation w...

Genotoxic effects of chlorophenoxy herbicide diclofop-methyl in mice in vivo and in human lymphocytes in vitro

Silene bolanthoides Quezel, Contandr. & Pamukc. is an endemic species from Kazdagi (Mt. Ida), Canakkale-Balikesir, Turkey. In order to develop an efficient shoot regeneration protocol, the leaf, nodal and internodal explants of S. bolanthoides were cultured on Murashige and Skoog (MS) medium containing benzyladenine (BA) alone or in combination with α-naphthaleneacetic acid (NAA). The highest number of regenerated shoots (5.75±0.1) was obtained from nodal explants that were cultured on MS medium with 8.8 µM BA+0.54 µM NAA. Regenerated shoots were rooted on MS medium without plant growth regulators (PGRs). Rooted plants (2-3 cm) were separately transferred to pots containing a mixture of peat and perlite (3:1 v/v) and acclimatized successfully in a growth chamber. Genetic stability of the propagated plants was assessed by flow cytometry and cytological analysis. Flow cytometry analysis demonstrated that regenerated plants had 2.61±0.01 pg nuclear DNA (2C) and seed-derived plants had on average 2.58±0.02 pg/2C. Cytological analysis showed that the regenerated plants had the same chromosome number as seed-derived plants of S. bolanthoides (2n=24). It was determined that regenerated plants were uniform in chromosome number and had a similar DNA content to the seed-derived ones, indicating that the described efficient shoot regeneration protocol can be applied for ex situ conservation of this species. https://doi.org/10.2298/ABS170410033C Received: April 10, 2017; Revised: August 9, 2017; Accepted: August 16, 2017; Published online: September 12, 2017 How to cite this article: Corduk N, Yucel G, Akinci N, Tuna M, Esen O. In vitro propagation of Silene bolanthoides Quezel, Contandr. & Pamukc. and assessment of genetic stability by flow cytometry. Arch Biol Sci. 2018;70(1):141-8.

/pdf/in-vitro-propagation-of-silene-bolanthoides-quezel-contandr-44dq96lm7f.pdf

In vitro propagation of Silene bolanthoides Quézel, Contandr. & Pamukç. and assessment of genetic stability by flow cytometry

In this study, flow cytometry and cytological analysis was used to evaluate the genetic stability of Digitalis trojana Ivanina plants regenerated via indirect shoot organogenesis. For in vitro propagation, leaf explants were excised from seedlings grown in sterile conditions and cultured MS medium supplemented with 3.0 mg/L BAP, 0.1 mg/L NAA. Shoots and calli were subcultured for a period of 2 weeks for shoot multiplication. For rooting, shoots were separated individually and transferred to MS medium containing 0.1% activated charcoal. Genetic stability of the regenerated plants was assessed by flow cytometry and cytological analyses. Flow cytometric analysis revealed that regenerated plantlets had as 2.82 pg nuclear DNA (2C) and seed-derived plants had on average 2.80 pg/2C. Cytological analysis showed that regenerated plantlets have the same number chromosome with seed-derived plantlets of D. trojana (2n=56). Our results have showed that the plantlets propagated in MS medium with 3 mg/L BAP, 0.1 mg/L NAA didn’t differ genetically from donor plants. Therefore, this system can be effective and suitable for clonal propagation of D. trojana . Our result also confirmed that flow cytometry is fast, easy, accurate and relatively cheap method to determine ploidy of in vitro propagated D. trojana plantlets.

/pdf/assessment-of-the-genetic-stability-of-indirect-shoot-2lwarz463e.pdf

Assessment of The Genetic Stability of Indirect Shoot Organogenesis-Derived Plantlets of Digitalis Trojana Ivanina by Flow Cytometry and Cytological Analyses

EVALUATION OF GENOTOXICITY AND CYTOTOXICITY OF DODINE (1-dodecylguanidium acetate) BY Allium TEST

This research was to evaluate the effects of dodine on the physiology of Vicia faba L. through total protein content and peroxidase (POX) activity . According to this, four weeks old seedlings were treated with 0.04 mL/L, 0.08 mL/L and 0.16 mL/L concentrations of dodine. Then, the leaves of plant samples were harvested with 24, 48 and 72 hours intervals. Our results have showed that while the total protein content were decreased significantly, POX activity was increased with the increased concentration of dodine and exposure time when compared with control seedlings. This research have shown that dodine can stimulate the plant defense system.

/pdf/dodin-fungusitinin-vicia-faba-l-bitkisinde-total-protein-avlnu12dxi.pdf

Dodin fungusitinin Vicia faba L. bitkisinde total protein miktarı ve peroksidaz aktivitesi üzerine etkisi

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