WUSCHEL controls genotype-dependent shoot regeneration capacity in potato.

Plant cells can reprogram their fate. The combinatorial actions of auxin and cytokinin dedifferentiate somatic cells to regenerate organs, which can develop into individual plants. As transgenic plants can be generated from genetically modified somatic cells through these processes, cell fate transition is an unavoidable step in crop genetic engineering. However, regeneration capacity closely depends on the genotype, and the molecular events underlying these variances remain elusive. In the present study, we demonstrated that WUSCHEL (WUS)-a homeodomain transcription factor-determines regeneration capacity in different potato (Solanum tuberosum) genotypes. Comparative analysis of shoot regeneration efficiency and expression of genes related to cell fate transition revealed that WUS expression coincided with regeneration rate in different potato genotypes. Moreover, in a high-efficiency genotype, WUS silencing suppressed shoot regeneration. Meanwhile, in a low-efficiency genotype, regeneration could be enhanced through the supplementation of a different type of cytokinin that promoted WUS expression. Computational modeling of cytokinin receptor-ligand interactions suggested that the docking pose of cytokinins mediated by hydrogen bonding with the core residues may be pivotal for WUS expression and shoot regeneration in potatoes. Furthermore, our whole-genome sequencing analysis revealed core sequence variations in the WUS promoters that differentiate low- and high-efficiency genotypes. The present study revealed that cytokinin responses, particularly WUS expression, determine shoot regeneration efficiency in different potato genotypes.

Assessing the combined toxic effects of metaldehyde mollucide.

The excessive use of metaldehyde in agriculture to combat mollusks endangers both the environment and non-target organisms. The aim of this study is to investigate the toxicity caused by metaldehyde in Allium cepa with the help of physiological, cytogenetic, biochemical and anatomical parameters. Also, DNA fragmentation caused by metaldehyde in root tip cells was measured by the "Comet Assay" method. The control group was germinated with tap water and the application groups were germinated with 20 mg/L metaldehyde, 40 mg/L metaldehyde, 100 mg/L metaldehyde and 200 mg/L metaldehyde for 72 h. The results of the physiological parameters showed that metaldehyde had a growth-limiting effect in A. cepa, depending on the application dose. According to root elongation levels, the EC50 (effective concentration) value for metaldehyde was 60.6 mg/L in A. cepa. As the treatment dose increased, the incidence of micronucleus and chromosomal aberrations gradually increased while mitotic index decreased. Metaldehyde exposure induced damages such as sticky chromosome, fragment, unequal distribution of chromatin, reverse polarization, bridge, and multipolar anaphase. In addition, metaldehyde caused cell damage in epidermis and cortex, thickening of the cortex cell wall and flattened cell nucleus in root meristem. Increasing doses of metaldehyde application also increased malondialdehyde levels, superoxide dismutase and catalase activities. As a result, it has been determined that the toxicity of metaldehyde in plants is versatile and the A. cepa test material is a suitable biological indicator to determine this toxicity.

Cyto-Genotoxic and Behavioral Effects of Flubendiamide in Allium cepa Root Cells, Drosophila melanogaster and Molecular Docking Studies.

Flubendiamide (FLB) is an insecticide that is commonly employed to control pests on a variety of vegetables and fruits, with low toxicity for non-target organisms. However, due to its widespread use, the environmental risks and food safety have become major concerns. In this study, the toxicity potential of FLB was studied in the model organisms, Allium cepa and Drosophila melanogaster. The cyto-genotoxic effects of FLB on the root growth, mitotic index (MI), chromosomal aberrations (CAs) and deoxyribonucleic acid (DNA) damage in A. cepa root meristematic cells were investigated using the root growth inhibition Allium test and Comet assays. FLB caused CAs in the form of disturbed ana-telophase, chromosome laggards, stickiness, anaphase-bridge and polyploidy depending on the concentration and the exposure time. The toxicity and genotoxicity of FLB at various doses (0.001, 0.01, 0.1 and 1 mM) on D. melanogaster were investigated from the point of view of larval weight and movement, pupal formation success, pupal position, emergence success and DNA damage, respectively. FLB exposure led to a significant reduction of the locomotor activity at the highest concentration. While DNA damage increased significantly in the FLB-treated onions depending on the concentration and time, DNA damage in the FLB-treated D. melanogaster significantly increased only at the highest dose compared to that which occurred in the control group. Moreover, to provide a mechanistic insight into the genotoxic and locomotion-disrupting effects of FLB, molecular docking simulations of this pesticide were performed against the DNA and diamondback moth (DBM) ryanodine receptor (RyR) Repeat34 domain. The docking studies revealed that FLB binds strongly to a DNA region that is rich in cytosine-guanine-adenine bases (C-G-A) in the minor groove, and it displayed a remarkable binding affinity against the DBM RyR Repeat34 domain.

Transcriptome-wide identification of NAC (no apical meristem/Arabidopsis transcription activation factor/cup-shaped cotyledon) transcription factors potentially involved in salt stress response in garlic.

Soil salinity has been an increasing problem worldwide endangering crop production and human food security. It is an ideal strategy to excavate stress resistant genes and develop salt tolerant crops. NAC (no apical meristem/Arabidopsis transcription activation factor/cup-shaped cotyledon) transcription factors have been demonstrated to be involved in salt stress response. However, relevant studies have not been observed in garlic, an important vegetable consumed in the world. In this study, a total of 46 AsNAC genes encoding NAC proteins were identified in garlic plant by transcriptome data. Phylogenetic analysis showed that the examined AsNAC proteins were clustered into 14 subgroups. Motif discovery revealed that the conserved domain region was mainly composed of five conserved subdomains. Most of the genes selected could be induced by salt stress in different tissues, indicating a potential role in salt stress response. Further studies may focus on the molecular mechanisms of the AsNAC genes in salt stress response. The results of the current work provided valuable resources for researchers aimed at developing salt tolerant crops.

Efficient micropropagation of Thunbergia coccinea Wall. and genetic homogeneity assessment through RAPD and ISSR markers.

Thunbergia coccinea Wall. ex D. Don being a rare, ornamental and medicinal plant of India, is needed to propagate for conserving the germplasm and analyzing its phytochemical compounds in the future. A reliable protocol for direct in vitro propagation using nodal shoot meristem of T. coccinea as explant was standardized. The highest number of shoots per explant (22.17 ± 0.54) with maximum shoot length (2.36 ± 0.28) in cm was obtained in Murashige and Skoog (MS) medium supplemented with 9.70 µM of 6-furfurylaminopurine (Kinetin) and 0.053 µM of α-naphthaleneacetic acid (NAA) combination, among all the different plant growth regulators (PGR's) and concentrations tested. The aforesaid PGR's combination was optimum for axillary shoot bud induction and multiplication in T. coccinea. The best rooting was observed on the half-strength MS medium fortified with 2.68 µM NAA with the highest number of roots per shoot (3.75 ± 0.12) and maximum length (5.22 ± 0.32) in cm. All the in vitro raised plantlets were acclimatized in sterile sand and soil mixture (1:1) with a survival rate of 70% on earthen pots under greenhouse conditions. PCR-based RAPD (Random Amplified Polymorphic DNA) and ISSR (Inter-Simple Sequence Repeat) molecular markers were employed to determine the genetic homogeneity amongst the plantlets. Twelve (12) RAPD and nine (9) ISSR primers developed a total of 104 and 91 scorable bands, respectively. The band profiles of micropropagated plantlets were monomorphic to the mother, donor in vivo plant, and similarity values varied from 0.9542-1.000. The dendrogram generated through UPGMA (unweighted pair group method with arithmetic mean) showed 99% similarities amongst all tested plants confirming the genetic uniformity of in vitro raised plants.

Chromosome behavior during meiosis in pollen mother cells from Saccharum officinarum × Erianthus arundinaceus F1 hybrids.

BACKGROUND: In recent years, sugarcane has attracted increasing attention as an energy crop. Wild resources are widely used to improve the narrow genetic base of sugarcane. However, the infertility of F1 hybrids between Saccharum officinarum (S. officinarum) and Erianthus arundinaceus (E. arundinaceus) has hindered sugarcane breeding efforts. To discover the cause of this infertility, we studied the hybridization process from a cytological perspective. RESULTS: We examined the meiotic process of pollen mother cells (PMCs) in three F1 hybrids between S. officinarum and E. arundinaceus. Cytological analysis showed that the male parents, Hainan 92-77 and Hainan 92-105, had normal meiosis. However, the meiosis process in F1 hybrids showed various abnormal phenomena, including lagging chromosomes, micronuclei, uneven segregation, chromosome bridges, and inability to form cell plates. Genomic in situ hybridization (GISH) showed unequal chromatin distribution during cell division. Interestingly, 96.70% of lagging chromosomes were from E. arundinaceus. Furthermore, fluorescence in situ hybridization (FISH) was performed using 45S rDNA and 5S rDNA as probes. Either 45S rDNA or 5S rDNA sites were lost during abnormal meiosis, and results of unequal chromosomal separation were also clearly observed in tetrads. CONCLUSIONS: Using cytogenetic analysis, a large number of meiotic abnormalities were observed in F1. GISH further confirmed that 96.70% of the lagging chromosomes were from E. arundinaceus. Chromosome loss was found by further investigation of repeat sequences. Our findings provide insight into sugarcane chromosome inheritance to aid innovation and utilization in sugarcane germplasm resources.

Evaluation of Cyto-genotoxicity of Perfluorooctane Sulfonate (PFOS) to Allium cepa.

Per- and polyfluoroalkyl substances (PFAS) have emerged as contaminants of global concern. Among several PFAS, perfluorooctanoic acid (PFOA) and perfluorooctane sulfonate (PFOS) are persistent and bioaccumulative compounds. We investigated the cyto-genotoxic potential of PFOS to Allium cepa root meristem cells. The A. cepa root tips were exposed to 6 different concentrations (1-100 mg L-1 ) of PFOS for 48 h. Reduction in mitotic index and chromosomal aberrations was measured as genotoxic endpoints in meristematic root cells. Exposure to PFOS significantly affected cell division by reducing the miotic index at higher concentrations (>10 mg L-1 ). The median effect concentration of PFOS to elicit cytotoxicity based on the mitotic index was 43.2 mg L-1 . Exposure to PFOS significantly increased chromosomal aberrations at concentrations >25 mg L-1 . The common aberrations were micronuclei, vagrant cells, and multipolar anaphase. The alkaline comet assay revealed a genotoxic potential of PFOS with increased tail DNA percentage at concentrations >25 mg L-1 . To our knowledge, this is the first study to report the cyto-genotoxic potential of PFOS in higher plants. Environ Toxicol Chem 2021;40:792-798. © 2020 SETAC.

Toxicity assessment of zinc sulfate: A commonly used compound.

Because zinc sulfate (ZnSO4) is widely used in many fields such as biomedicine, electronics, and chemistry, it is important to evaluate its toxic effects. In this study, the cyto-genotoxic effects of ZnSO4 on meristematic cells in the root tip of Allium cepa L. were investigated. After calculating the effective concentration (EC50 = 70 ppm) of ZnSO4, A. cepa root tip cells were suspended for 24, 48, 72, and 96 h in solutions of 35 ppm (EC50/2), 70 ppm (EC50), and 140 ppm (EC50 × 2) concentrations. Using the counts of dividing cells, the mitotic index (MI) was calculated. Chromosome aberration index (CAI) was determined from percentages of abnormal cells. When the obtained data were statistically evaluated, it was determined that all application concentrations caused a significant decrease in MI and an increase in CAI compared to the control group (distilled water). It was concluded that increased ZnSO4 dose concentrations and exposure times caused cytotoxicity and genotoxicity in the root cells of A. cepa L.

Changes in gene expression in potato meristems treated with the sprout suppressor 1,4-dimethylnaphthalene are dependent on tuber age and dormancy status.

Commercial storage of potatoes often relies on the use of sprout inhibitors to prolong storage and reduce spoilage. The compound 1,4-dimethylnaphthalene (DMN) has seen increase application as a sprout inhibitor in the potato industry as older chemistries are being phased out. The mode of action of DMN is poorly understood as is the sensitivity of potato tissues to this new class of inhibitor. During storage potato tubers transition from a state of endo-dormant to eco-dormant and it is not known if the DMN response is consistent across this developmental transition. RNA-seq gene expression profiling was used to establish if stored potato tubers (Solanum tuberosum cv La Chipper) have differential sensitivity to DMN as tubers age. DMN was applied at three different times during storage; just after harvest when tubers are in endo-dormancy, midwinter at early eco-dormancy, and in spring during late eco-dormancy when sprouting was prevented via exposure to cold storage temperatures. Changes in gene expression were lowest during endo-dormancy while midwinter and spring treatments exhibited a greater and more diverse expression response. Functional analysis of differential gene expression demonstrated gene sets associated with DNA replication, cell division, and DNA methylation are suppressed after DMN treatment. However, gene sets associated with salicylic acid, jasmonic acid, abiotic and biotic stress responses are elevated by DMN only after endodormancy terminates. Gene clusters associated with pathogenesis related proteins PR-4 and PR-5 are also upregulated in response to DMN. These results indicate that DMN sensitivity changes as potato tubers age and transition from endo-dormant to eco-dormant in storage and the overall response is a shift in gene classes that regulate growth and response to stress.

Cytotoxic and Genotoxic Assessment of Silicon Dioxide Nanoparticles by Allium and Comet Tests.

Silicon nanoparticles gained a great interest due to its use in biomedical research. It is considered as safe and has been used in nanomedicine. But literature still states its toxicity depending upon the size and dose of silicon nanoparticles. So, current study was aimed to evaluate the cytotoxicity and genotoxicity of silicon dioxide nanoparticles (SiO2NPs) by Allium anaphase-telophase and Comet tests. Characterization of SiO2NPs showed the particle size as 16.12 ± 3.07 nm. The mean diameter of SiO2NPs was having range of 404.66 ± 93.39 nm in solution. Highest total anomalies (18.80 ± 0.45) were observed at 100 µg/mL, whereas least (11.2 ± 0.84) were observed by the 12.5 µg/mL concentration. There was concentration-response association in increased CAs and DNA damage. The highest concentration (100 µg/mL) of SiO2NPs induced the significant DNA damage (149.67 ± 1.15), whereas the least was observed by the negative control (2.67 ± 0.58). The current study revealed the cytotoxic and genotoxic effects of SiO2NPs on the root meristem cells of A. cepa.

Comparative cyto- and genotoxicity of 900 MHz and 1800 MHz electromagnetic field radiations in root meristems of Allium cepa.

In the last few decades, tremendous increase in the use of wireless electronic gadgets, particularly the cell phones, has significantly enhanced the levels of electromagnetic field radiations (EMF-r) in the environment. Therefore, it is pertinent to study the effect of these radiations on biological systems including plants. We investigated comparative cytotoxic and DNA damaging effects of 900 and 1800 MHz EMF-r in Allium cepa (onion) root meristematic cells in terms of mitotic index (MI), chromosomal aberrations (CAs) and single cell gel electrophoresis (comet assay). Onion bulbs were subjected to 900 and 1800 MHz (at power densities 261 ±â€¯8.50 mW m-2 and 332 ±â€¯10.36 mW m-2, respectively) of EMF-r for 0.5 h, 1 h, 2 h, and 4 h. Root length declined by 13.2% and 12.3%, whereas root thickness was increased by 46.7% and 48.3% after 4 h exposure to 900 MHz and 1800 MHz, respectively. Cytogenetic studies exhibited clastogenic effect of EMF-r as depicted by increased CAs and MI. MI increased by 36% and 53% after 2 and 4 h exposure to 900 MHz EMF-r, whereas it increased by 41% and 67% in response to 1800 MHz EMF-r. Aberration index was increased by 41%-266% and 14%-257% during 0.5-4 h of exposure to 900 MHz and 1800 MHz, respectively, over the control. EMF-r exposure decreased % head DNA (DNAH) and increased % tail DNA (DNAT) and olive tail moment (OTM) at both 900 and 1800 EMF-r. In 4 h exposure treatments, head DNA (%) declined by 19% and 23% at 900 MHz and 1800 MHz, respectively. DNAT and OTM were increased by 2.3 and 3.7 fold upon exposure to 900 MHz EMF-r over that in the control, whereas 2.8 and 5.8 fold increase was observed in response to 1800 MHz EMF-r exposure for 4 h and the difference was statistically significant. The study concludes that EMF-r in the communication range (900 and 1800 MHz) adversely affect root meristems in plants and induce cytotoxic and DNA damage. EMF-r induced DNA damage was more pronounced at 1800 MHz than that at 900 MHz.

Global epigenetic changes of histone modification under environmental stresses in rice root.

Abiotic stresses are non-living factors with negative morphological and physiological effects on living organisms. Substantial evidence exists that gene expression changes during plant cell growth are regulated by chromatin reconfiguration and histone modification. Several types of histone modifications are dramatically transformed in stress-responsive gene regions under drought stress conditions. Environmental stresses also cause the root apical meristem (RAM) region to decelerate root growth. In this study, we investigated how quantitative changes in epigenetic markers in this region influence rice morphology and physiology. Both iron and salinity treatments changed the epigenetic landscape from euchromatic to heterochromatic according to heterochromatin (H3K9me2) and euchromatin (H3K4me) markers, especially in the proximal meristem region. Moreover, supplementation with external abscisic acid (ABA) was able to mimic the effect of environmental stresses on global epigenetic changes. In contrast, the addition of external auxin (IAA) to rice under saline conditions affected heterochromatin formation without influencing euchromatin transformation. Chromatin dynamics is therefore believed to be directly connected to plant growth regulator signaling. We discuss insights into the role of plant growth regulators: ABA and IAA, peroxide signaling, and their effects on the global epigenetic change of histone modification under abiotic stresses.

The transcriptome of potato tuber phellogen reveals cellular functions of cork cambium and genes involved in periderm formation and maturation.

The periderm is a protective corky tissue that is formed through the cambial activity of phellogen cells, when the outer epidermis is damaged. Timely periderm formation is critical to prevent pathogen invasion and water loss. The outer layers of the potato periderm, the tuber skin, serves as a model to study cork development. Early in tuber development the phellogen becomes active and produces the skin. During tuber maturation it becomes inactive and the skin adheres to the tuber flesh. The characterization of potato phellogen may contribute to the management of costly agricultural problems related to incomplete skin-set and the resulting skinning injuries, and provide us with new knowledge regarding cork development in planta. A transcriptome of potato tuber phellogen isolated by laser capture microdissection indicated similarity to vascular cambium and the cork from trees. Highly expressed genes and transcription factors indicated that phellogen activation involves cytokinesis and gene reprograming for the establishment of a dedifferentiation state; whereas inactivation is characterized by activity of genes that direct organ identity in meristem and cell-wall modifications. The expression of selected genes was analyzed using qPCR in native and wound periderm at distinct developmental stages. This allowed the identification of genes involved in periderm formation and maturation.

Mitogen-activated protein kinase 6 integrates phosphate and iron responses for indeterminate root growth in Arabidopsis thaliana.

MAIN CONCLUSION: A MAPK module, of which MPK6 kinase is an important component, is involved in the coordination of the responses to Pi and Fe in the primary root meristem of Arabidopsis thaliana. Phosphate (Pi) deficiency induces determinate primary root growth in Arabidopsis through cessation of cell division in the meristem, which is linked to an increased iron (Fe) accumulation. Here, we show that Mitogen-Activated Protein Kinase6 (MPK6) has a role in Arabidopsis primary root growth under low Pi stress. MPK6 activity is induced in roots in response to low Pi, and such induction is enhanced by Fe supplementation, suggesting an MPK6 role in coordinating Pi/Fe balance in mediating root growth. The differentiation of the root meristem induced by low Pi levels correlates with altered expression of auxin-inducible genes and auxin transporter levels via MPK6. Our results indicate a critical role of the MPK6 kinase in coordinating meristem cell activity to Pi and Fe availability for proper primary root growth.

Toxicological effects of comercial formulations of fungicides based on procymidone and iprodione in seedlings and root tip cells of Allium cepa.

In this study the phytotoxic, cytotoxic, genotoxic and mutagenic effects of two commercial fungicide-active compounds, procymidone (PR) and iprodione (IP), were determined. The parameters evaluated were germination and root growth, mitotic index, chromosomal and nuclear aberrations, and molecular analyses were also performed in the model plant Allium cepa L. The results demonstrated that the active compounds PR and IP were phytotoxic, delaying germination and slowing the development of A. cepa seedlings. Moreover, PR and IP showed cytogenotoxicity towards A. cepa meristematic cells, inducing chromosomal changes and cell death. The mutagenic activity of the active compounds was demonstrated by the detection of DNA changes in simple sequence repeat (SSR) and inter-simple sequence repeat (ISSR) markers in the treated cells compared to the negative control. Together, these results contribute to a better understanding of the damage caused by these substances in living organisms and reveal a promising strategy for prospective studies of the toxic effects of environmental pollutants.

Sex change in kiwifruit (Actinidia chinensis Planch.): a developmental framework for the bisexual to unisexual floral transition.

KEY MESSAGE: The developmental morphology of male and female kiwifruit flowers is tracked to delimit a framework of events to aid the study of divergence in floral gene expression. The transition from hermaphrodite to unisexual development of kiwifruit (Actinidia chinensis Planch) flowers has been reported previously, but differences in gene expression controlling sexual development for this species have not been associated with the major developmental changes occurring within pistils. We investigated the key stages in male and female flower development to define the point at which meristematic activities diverge in the two sexes. A combination of scanning electron microscopy and light microscopy was used to investigate pistil development from the earliest stages. We identified seven distinct stages characterized by differences in ovary size and shape, macrosporogenesis, ovule primordium development, anther locule lengthening, microspore wall thickening, and pollen degeneration. Sex differences were evident from the initial stage of development, with a laterally compacted gynoecium in male flowers. However, the key developmental stage, at which tissue differentiation clearly deviated between the two sexes, was stage 3, when flowers were 3.5 to 4.5 mm in length at approximately 10 d from initiation of stamen development. At this stage, male flowers lacked evident carpel meristem development as denoted by a lack of ovule primordium formation. Pollen degeneration in female flowers, probably driven by programmed cell death, occurred at the late stage 6, while the final stage 7 was represented by pollen release. As the seven developmental stages are associated with specific morphological differences, including flower size, the scheme suggested here can provide the required framework for the future study of gene expression during the regulation of flower development in this crop species.

Functional Analysis of Coilin in Virus Resistance and Stress Tolerance of Potato Solanum tuberosum using CRISPR-Cas9 Editing.

The role of the nuclear protein coilin in the mechanisms of resistance of potato Solanum tuberosum cultivar Chicago to biotic and abiotic stresses was studied using the CRISPR-Cas9 technology. For the coilin gene editing, a complex consisting of the Cas9 endonuclease and a short guide RNA was immobilized on gold or chitosan microparticles and delivered into apical meristem cells by bioballistics or vacuum infiltration methods, respectively. Editing at least one allele of the coilin gene considerably increased the resistance of the edited lines to infection with the potato virus Y and their tolerance to salt and osmotic stress.

Phytotoxicity, cytotoxicity and genotoxicity evaluation of organic and inorganic pollutants rich tannery wastewater from a Common Effluent Treatment Plant (CETP) in Unnao district, India using Vigna radiata and Allium cepa.

The leather industry is a major source of environmental pollution in India. The wastewater generated by leather industries contains very high pollution parameters due to the presence of a complex mixture of organic and inorganic pollutants even after the treatment at a Common Effluent Treatment Plant (CETP) and disturbs the ecological flora and fauna. The nature, characteristics and toxicity of CETP treated wastewater is yet to be fully elucidated. Thus, this study aims to characterize and evaluate the toxicity of CETP treated tannery wastewater collected from the Unnao district of Uttar Pradesh, India. In addition to measuring the physico-chemical parameters, the residual organic pollutants was identified by GC-MS analysis and phytotoxicity, cytotoxicity and genotoxicity of the treated wastewater was evaluated using Vigna radiata L. and Allium cepa L. Results showed that the treated wastewater contained very high pollution parameters (TDS 3850 mg/L, BOD 680 mg/L, COD-1300 mg/L). GC-MS analysis revealed the presence of various types of residual organic pollutants including benzoic acid, 3-[4,-(T-butyl) Phenyl] furan-2-5-dione, benzeneacetamide, resorcinol, dibutyl phthalate, and benzene-1,2,4-triol. Further, toxicological studies showed the phytotoxic nature of the wastewater as it inhibited seed germination in V. radiata L. and root growth of A. cepa. Genotoxicity was evidenced in the root tip cell of A. cepa where chromosomal aberrations (stickiness, chromosome loss, C-mitosis, and vagrant chromosome) and nuclear abnormalities like micronucleated and binucleated cells were observed. Thus, results suggested that it is not safe to discharge these wastewater into the environment.

Target-mimicry based diminution of miRNA167 reinforced flowering-time phenotypes in tobacco via spatial-transcriptional biases of flowering-associated miRNAs.

Evolutionarily conserved microRNAs such as miR156, miR159, miR167 and miR172 tightly regulate the extensive array of gene expression during flowering in plants, through instant and long-term alterations in the expression of their target genes. Here we employed a novel target-mimicry approach for the diminution of auxin signalling regulator miRNA167 by developing mimic-transgenic lines in tobacco, to investigate the transcriptional biases of flowering-associated miRNAs in apical and floral meristematic tissues and their phenotypic implications. Recorded morpho-alterations such as uneven flowering-time phenotypes, anomalous floral organ formation, and large variations in the seed forming characteristics permitted us to determine the consequence of the extent of miR167 expression diminution accompanying the transcriptional biases of interrelated miRNAs. We demonstrate that percent diminution of miR167 gene expression is proportionally associated with both early and late flowering-time phenotypes in mimic lines. Also, the associated miRNAs, miR156, miR159, and miR172 showed >90% transcriptional diminution in at least 'early-flowering' miR167 mimic lines. On contrary, low percentages of their respective diminution were recorded in 'late-flowering' lines. Evidently, the misexpression of miR156, miR159, and miR172 led to the over-expression of their respective target genes SPL9, AtMYB33-like and AP2 genes in mimic lines which resulted in assorted phenotypes. We describe the scope of spatial regulation of these microRNAs in floral bud tissues of mimic lines which showed negative- or very low (<25%) misexpression levels in early/late-flowering lines highlighting their roles in the acquisition of flowering mechanism. To our knowledge, this study represents the first characterization of transcriptional biases of flowering associated miRNAs in miR167-mimic lines and certainly augments our understanding of the importance of microRNA-mediated regulation of flowering in plants.

Cytogenetic and genotoxic effects of 2-chlorophenol on Allium cepa L. root meristem cells.

2-Chlorophenol (2-CP), a class of chlorinated organic pollutants like other chlorophenols, is used as intermediate in the synthesis of the higher chlorinated congeners, certain dyes, preservatives, herbicides, fungicides, and plastics. In this study, cytotoxic and genotoxic effects of 2-CP were investigated on the root meristem cells of Allium cepa for its effects on root growth, mitotic index (MI), mitotic phases, chromosomal abnormalities (CAs), and DNA damage by using Allium anaphase-telophase and Comet assays. EC50 of 2-CP value was determined as approximately 25 mg/L by Allium root growth inhibition test. Three concentrations of 2-CP (12.5, 25, and 50 mg/L), distilled water (negative control), and methyl methane sulfonate (MMS, 10 mg/L, positive control) were applied to onion stem cells under different exposure periods (24, 48, 72, and 96 h). All the applied doses of 2-CP slightly decreased MIs. 2-CP induced total CAs such as disturbed anaphase-telophase, chromosome laggards, stickiness, and bridges and also DNA damage at significant levels. These results demonstrate that 2-CP has genotoxic effects in A. cepa root meristematic cells.