To understand better the plant response to ozone, we isolated and characterized an ozone-sensitive (ozs1) mutant strain from a set of T-DNA-tagged Arabidopsis thaliana ecotype Columbia. The mutant plants show enhanced sensitivity to ozone, desiccation and sulfur dioxide, but have normal sensitivity to hydrogen peroxide, low temperature and high light levels. The T-DNA was inserted at a single locus which is linked to ozone sensitivity. Identification of the genomic sequences flanking the T-DNA insertion revealed disruption of a gene encoding a transporter-like protein of the tellurite resistance/C(4)-dicarboxylate transporter family. Plants with either of two different T-DNA insertions in this gene were also sensitive to ozone, and these plants failed to complement ozs1. Transpiration levels, stomatal conductance levels and the size of stomatal apertures were greater in ozs1 mutant plants than in the wild type. The stomatal apertures of ozs1 mutant plants responded to light fluctuations but were always larger than those of the wild-type plants under the same conditions. The stomata of the mutant and wild-type plants responded similarly to stimuli such as light, abscisic acid, high concentrations of carbon dioxide and ozone. These results suggest that OZS1 helps to close stomata, being not involved in the responses to these signals.
This study explained about machining parameters of Al5086/Flyash/Sic hybrid metal matrix composites by the Taguchi technique. Al5086 reinforced in SiC (5–10 wt %) and 8% weight of flyash are retained as constants. The specimens are prepared with the help of the stir casting method. The material removal rate was examined by electrochemical machining under various parameters such as feed rate (0.15–0.30 mm/min), voltage (10–20 V), and electrolyte concentration (20–35 g/litre). Taguchi’s L16 orthogonal array was selected for design of experiments (DOEs), and 16 experimental tests were conducted to examine the effect of the selected machining parameters employed to identify the best optimal levels and also to investigate the effect of electrochemical machining parameters on MRR determined by Minitab-18.
An ozone-sensitive mutant was isolated from T-DNA-tagged lines of Arabidopsis thaliana. The T-DNA was inserted at a locus on chromosome 3, where two genes encoding glycolate oxidases, GOX1 and GOX2, peroxisomal enzymes involved in photorespiration, reside contiguously. The amounts of the mutant's foliar transcripts for these genes were reduced, and glycolate oxidase activity was approximately 60% of that of the wild-type plants. No difference in growth and appearance was observed between the mutant and the wild-type plants under normal conditions with ambient air under a light intensity of 100 µmol photons m-2 s-1. However, signs of severe damage, such as chlorosis and ion leakage from the tissue, rapidly appeared in mutant leaves in response to ozone treatment at a concentration of 0.2 µl l-1 under a higher light intensity of 350 µmol photons m-2 s-1 that caused no such symptoms in the wild-type plant. The mutant also exhibited sensitivity to sulfur dioxide and long-term high-intensity light. Arabidopsis mutants with deficiencies in other photorespiratory enzymes such as glutamate:glyoxylate aminotransferase and hydroxypyruvate reductase also exhibited ozone sensitivities. Therefore, photorespiration appears to be involved in protection against photooxidative stress caused by ozone and other abiotic factors under high-intensity light.
In general, antimicrobial agents are often used in micropropagation techniques to obtain contaminant free clones. The objective of the present study was to evaluate the effects of bavistin and cefotaxime on producing contaminant free plants of Ruellia tuberosa cultured on MS supplemented with phytohormones. Field grown nodal explants of Ruellia tuberosa was used to regenerate entire plants via direct organogenesis. Among the decontaminants tested, the fungicide bavistin along with higher concentration of BAP (2.0 mg/l) and lower concentration of NAA (1.0 mg/l) was the most effective in regeneration and producing contaminant free shoots from cultured explants. This fungicide at 300 mg/l minimised fungal contamination with survival rate of 54%. While the addition of decontaminant cefotaxime at low concentration (200 mg/l) along with same concentration of BAP and NAA stimulated the bud formation and controlled the bacterial contamination. However, its increasing concentration adversely affected the survival rate of Ruellia tuberosa. These findings clearly showed that low concentrations of bavistin and cefotaxime were not only non-toxic but also facilitated bud regeneration. The results achieved showed the decisive role not only of the use of successful fungicides and antibiotics, but also of their sufficient doses were very important in reducing contamination and helping multiple shoot proliferation.
Plant Tissue Cult. & Biotech. 31(1): 1-12, 2021 (June)
An efficient protocol was standardized for successful regeneration of Cissampelos pareira (L.) through indirect organogenesis. Nodal explants were cultured on MS fortified with 0.5 ± 1.0 mg/l BAP, Kn either single or in combination with NAA 0.5 mg/l. The combinations induced profuse, compact, light green to greenish coloured calli. Some differences in the morphology of callus such as change in the colour and texture was also observed with increasing the concentration of BAP 0.5 ± 2.0 mg/l + NAA 0.5 mg/l. Maximum callus induction was observed on 1.0 mg/l BAP and 0.5 mg/l NAA showed greenish, friable and granular lush colour. The calli were subcultured on fresh MS that contained BAP and Kn single or in combination with NAA (BAP 0.5 ± 2.0 mg/l, Kn 0.5 ± 2.0 mg/l, NAA 0.5 mg/l). The maximum regeneration frequency of shoot organogenesis was recorded on BAP (2.0 mg/l) + NAA (0.5 mg/l). Healthy microshoots were separated and transferred to the rooting medium. Here, MS augmented with IBA 1.0 mg/l showed maximum rooting. Well rooted plantlets were transferred to the field and maximum survival frequency was recorded when BAP (1.0 mg/l) + NAA (0.5 mg/l) for callus induction, for shooting BAP (2.0 mg/l) + NAA (0.5 mg/l) and for rooting IBA (1.0 mg/l) was used. The regenerated whole plants were subjected for hardening where the maximum survival frequency was found to be 80%. This reproducible protocol can be used for regeneration and genetic transformation studies.
Axillary bud explants of Oxalis corniculta grown in the field were excised and inoculated on MS medium with various concentrations and combinations of auxins (2, 4-D and NAA) and cytokinins (BAP and Kn) for indirect shoot formation through callus induction. Indirect organogenesis was achieved by the application of PGRs such as BAP, Kn, and NAA in the culture medium. The maximum mean shoot number (9.2) and regeneration frequency (77%) was found with BAP (2.0 mg/l) and NAA (0.5 mg/l). Healthy microshoots were separated and transferred to rooting medium supplemented with NAA, IBA and IAA. MS medium augmented with NAA 2.0 mg/l was found to be best for rooting. The regenerated plantlets were then hardened, acclimatized and where exhibited 75% survivability. The reproducible protocol can be used for regeneration and genetic transformation studies.
Plant Tissue Cult. & Biotech. 32(2): 181-191, 2022 (December)
Background: Optimization of co-cultivation parameters during Agrobacterium-mediated transformation to Euphorbia tirucalli evaluated were bacterial density, infection period, acetosyringone (AS) concentration and co-cultivation temperature. Optimized parameters resulted in high transformation efficiency 3 fold increase at transient GUS expression .The optimized conditions were Agrobacterium tumefaciens growth phase of A600nm 0.17, infection period of 30 min, addition of acetosyringone (AS) in co-cultivation medium (100 µM) and cocultivation temperature of 25°C.
Methods: Nodal explants were used for transformation. Bacterial culture was added to 50 ml of liquid YEP medium with kanamycin and rifampicin and grown until reaching the growth phase (A600nm). Bacterial density ranged from A600nm 0.17, 0.56, 0.8 and 1.2 OD were used in the present study. The co-cultivation medium made of solid MS medium consisted of NAA 2 mg L-1 and various concentrations of AS at 0, 50, 100, 200, 400, 600 and 800 µM. Histochemical analysis of gus gene expression was carried out.
Result: Higher bacterial density resulted in more transformation efficiency, but also higher necrosis in the explants. Dilution of bacterial suspension reduced necrosis in explants and resulted in higher transformation. The transformation efficiency is 72% when the infection process was carried out with acetosyringone in co-cultivation medium (100 µM). Our studies proved that among the optimized conditions, cocultivation temperature and acetosyringone were the critical parameters during Agrobacterium mediated transformation.
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