Determination of genotoxic damages of picloram and dicamba with comet assay in Allium cepa rooted in tissue culture and distilled water.

BACKGROUND: Many genotoxicity tests allow us to understand the mechanism of damages on genetic material occurring in living organisms against various physical and chemical agents. One of them is the Comet test. The current study aimed to evaluate genotoxic caused by picloram and dicamba to root meristems of Allium cepa utilizing comet assay. METHODS: Two different protocols were used for rooting and auxin/pesticide application. (i) A. cepa bulbs were rooted in MS medium and then treated with Murashige and Skoog (MS) medium (control) and 0.67, 1.34, 2.01, 2.68, 3.35, 4.02, and 8.04 mg/L of picloram and dicamba using aseptic tissue culture techniques. (ii) A. cepa bulbs were then rooted in bidistilled water and treated with 0 (control), 0.67, 1.34, 2.01, 2.68, 3.35, 4.02, and 8.04 mg/L of picloram and dicamba in distilled water. The A. cepa root tip cells in both treatment groups were examined using comet test to find the possible DNA damaging effects of picloram and dicamba. RESULTS: The results obtained at all the concentrations were statistically compared with their control groups. Almost at all the concentrations of Picloram and dicamba increased comet tail intensity (%) and tail moment in roots treated in MS medium. Two highest concentrations revealed toxic effect. On the other hand, DNA damaging effect of both auxins was only noted on the highest (> 4.02 mg/L) in roots treated in distilled water. CONCLUSIONS: This study approve and confirm genotoxic effects of how growth regulators on plants. These findings give an evidence of DNA damage in A. cepa. Therefore, both picloram and dicamba should only be used in appropriate and recommended concentrations in agriculture to conserve ecosystem and to pose minimum threat to life.

Does 2,4-dichlorophenoxyacetic acid (2,4-D) induce genotoxic effects in tissue cultured Allium roots?

2,4-Dichlorophenoxyacetic acid (2,4-D) is a synthetic plant growth regulator that is highly toxic to most broad leaved plants and relatively nontoxic to monocotyledonous plants; is frequently used as weed killer. The study aimed to investigate cytogenetic effects of different concentrations of 2,4-D (0.67, 1.34, 2.01, 2.68, 3.35 and 4.02 mg/L) on Allium cepa bulblets' root tips treated for 24 and 48 h. The results showed six types of structural aberrations: C-mitosis, stickiness, laggards, bridges, fragments and multipolarity that varied numerically compared to control. It significantly affected mitotic index (MI) at 24 and 48 h treatment. In the Allium test, MI increased significantly at three lower concentrations (0.67, 1.34, 2.01 mg/L) after treatment with 2,4-D for 24 h and decreased significantly at higher concentration. Whereas, 2,4-D treatment for 48 h increased MI at all concentrations with significantly decreased MI at the highest concentration. The experiment was extended using comet test that did not reveal significant difference among treatments except for application of 4.02 mg/L 2,4-D for 48 h; where cell damages were verified by comet test. Rest of the concentrations for any duration of time were not damaging and toxic to cells. The results showed, visible mitodepressive action of 4.02 mg/L 2,4-D when treated for 48 h that had tendency to become toxic if the roots had been in touch with 2,4-D for a longer time.

Factors affecting efficient in vitro micropropagation of Muscari muscarimi Medikus using twin bulb scale.

Endemic Muscari muscarimi Medikus is the most fragrant plant among Muscari species and has a high ornamental potential. The natural populations of M. muscarimi, are severely affected by increased environmental pollution and urbanization. There is a need to develop a micropropagation method that should serve effectively for commercial propagation and conservation. Therefore, the study targeted to set up a strategy for efficient in vitro bulblet regeneration system of M. muscarimi using twin scale bulb explants on 1.0 × MS medium containing 4.44, 8.88, 17.76 µM BAP (6-Benzylaminopurine) plus 2.685, 5.37, 10.74 µM NAA (α-Naphthalene acetic acid). Maximum number of 19 daughter axillary bulblets and 16 daughter adventitious bulblets per twin bulb scale explant was regenerated on 1.0 × MS medium containing 17.76 µM BAP plus 10.74 µM NAA and 17.76 µM BAP plus 2.685 µM NAA respectively. The daughter bulblets regenerated on twin bulb scales on 8 out of 9 regeneration treatment could be easily rooted on 1.0 × MS medium containing 4.9 µM IBA (Indole-3-butyric acid). The daughter bulblets regenerated on 9th treatment (1.0 × MS medium containing 17.76 µM BAP plus 10.74 µM NAA) were transferred to 1.0 × MS medium containing 30 g/l sucrose to break negative carry over effect of this dose of BAP-NAA, where they grew 2-3 roots of variable length. Daughter bulblet diameter was increased by culturing them on 1.0 × MS medium containing 4.44 µM BAP plus 5.37 µM NAA. The results verified that both age and the source of explants had significant effect on regeneration. In another set of experiments, twin scales were obtained from in vitro regenerated daughter bulblets, although they induced bulblets, yet their bulblet regeneration percentage, mean number of bulblets per explant and their diameter were significantly reduced. In vitro regenerated bulblets were acclimatized in growth chamber under ambient conditions of temperature and humidity on peat moss, where they flowered. The study provides important information about selection of suitable micropropagation medium, strategies to improve bulblet diameter and rooting of M. muscarimi which offers a scope for commercial propagation.

Factors affecting in vitro plant regeneration of the critically endangered Mediterranean knapweed (Centaurea tchihatcheffii Fisch et. Mey).

Habitat destruction has resulted in the extinction of many plant species from the earth, and many more face extinction. Likely, the annual endemic Mediterranean knapweed (Centaurea tchihatcheffii) growing in the Golbasi district of Ankara, Turkey is facing extinction and needs urgent conservation. Plant tissue culture, a potentially useful technique for ex situ multiplication, was used for the restoration of this ill-fated plant through seed germination, micropropagation from stem nodes, and adventitious shoot regeneration from immature zygotic embryos. The seeds were highly dormant and very difficult to germinate. No results were obtained from the micropropagation of stem nodes. However, immature zygotic embryos showed the highest adventitious shoot regeneration on Murashige and Skoog (MS) medium, containing 1 mg l(-1) kinetin and 0.25 mg l(-1) NAA. Regenerated shoots were best rooted on MS medium containing 1 mg l(-1) IBA and transferred to the greenhouse for flowering and seed set. As such, the present work is the first record of in vitro propagation of critically endangered C. tchihatcheffii, using immature zygotic embryos, and is a step forward towards conservation of this indigenous species.