Due to the rapid increase of the world population, a large part of the existing agricultural lands was occupied by industrial and residential areas. Both conventional farming practices and plant biotechnology are used to obtain more yield from the available land per unit area. In both productions, chemicals called plant growth regulators (PGR) are used to influence plant growth and development such as repressing shoot growth, increasing branching, increasing blooming, detaching excess fruit, or modifying fruit maturity . Auxins are among the most widely used groups of PGRs. They are used in plant tissue culture, especially for callus induction and tissue and organ differentiation. They are available in both natural and synthetic forms. Dicamba, 2,4-D, and Picloram are two classes of synthetic auxins. They tend to exhibit higher activity compared to other auxins like NAA (1-Naphthaleneacetic acid), IAA (3-Indoleacetic acid), and IBA (Indole-3-butyric acid), promoting somatic embryo development in tissue culture . In the last decade, loss of the 30% of the agricultural production in the world has resulted from agricultural pests; about 13% of these are weeds . Picloram and Dicamba are commonly used auxin herbicides in combating weeds that cause high crop loss each year, competing with cultivated crops in terms of nutrients, water, light, and location they take from the soil .
Picloram or 4-amino-3,5,6-trichloropicolinic acid is also included in the class of picloram picolinic acid herbicides or pyridine herbicides due to its herbicidal action . Picloram usage comes after glyphosate, throughout the world . This auxin herbicide is commonly used to kill dicotyledonous weeds in crop fields and pastures . According to the data described in the EPA (1995) report, Picloram does not cause chromosomal damage; declared non-toxic at low doses to birds, mammals, and aquatic species. In high concentrations, it has been reported to cause vascular tissue damage and inhibit cell division .
Dicamba or 3,6-dichloro-2-methoxybenzoic acid, on the other hand, is a systemic broad-spectrum herbicide from benzoic acid derivative synthetic auxins and is mostly used in agricultural areas to control single and perennial weeds. Besides, there are many commercial and industrial applications such as the maintenance of pasture and forest areas, cleaning of golf courses, and parks-gardens in residential areas from weeds . With the development of Monsanto's dicamba-resistant GM soybeans and cotton, dependence on dicamba has increased in recent years , which means that concentrations on non-target crops and in the environment will increase steadily . While the use of auxin herbicides provides a great advantage in killing harmful plants, they are common causes of soil and water pollution . Furthermore, many macro and microorganisms with their residues in the soil, they can be washed from the soil and directly reach groundwater and surface water such as rivers, lakes, and sea , or accumulate in organisms living there and negatively affect the food chain [12, 13]. Samanic et al.  noted that farmers in agricultural areas, where dicamba was applied, were more frequently caught in lung and colon cancer than other individuals, according to the agricultural health study data. Similarly, Lerro et al.  revealed a relationship between the usage of dicamba and many types of cancer based on the data of agricultural health studies conducted in the last twelve years. These results reveal that exposure of living organisms to chemical agents such as herbicides can cause damage to their genetic material.
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 or SCGE-Single Cell Gel Electrophoresis that was firstly used in 1984 by Östling and Johanson to evaluate DNA damage. Comet technique is a fast, reliable, and cost-effective method for observing DNA damage . It is currently used to directly determine DNA damages and repair in both animal and plant materials [17–22]. There are several model species like Arabidopsis thaliana, Vicia faba, Nicotiana tabacum or Allium cepa to perform Comet test [20, 23]. Allium cepa is the most widely preferred plant species and has been used in genotoxicity studies related to many chemical agents [19, 21, 24, 25]. When the literature was searched, it was seen that there were only a few studies conducted in recent years on the DNA damaging effect of picloram in plants. In one of these studies, Taspinar et al.  have revealed changes in DNA methylation using CREDRA (Coupled Restriction Enzyme Digestion Random Amplification) techniques in bean (Phaseolus vulgaris) and DNA damage using RAPD (Random Amplified Polymorphic DNA) techniques. Mohammed and Ma  have demonstrated the genotoxic effect of picloram using the micronucleus test on Tradescantia.
RAPD and CREDRA methods were applied to test the effects of picloram and dicamba, respectively, on genetic and epigenetic changes in mature embryo culture of barley (Hordeum vulgare L.). While authors have observed DNA hypomethylation following dicamba treatment, hypermethylation of DNA were confirmed utilizing higher doses of picloram . However, as far as we know, no study has been found investigating the genotoxic potential of picloram using the comet test.
A few studies were conducted using different genotoxicity tests to check the effects of Dicamba on nontarget plants. For example, Abass et al.  have examined the genotoxic effects of Dicamba on the Hillawii variety of palm (Phoenix dactylifera L.) plant in tissue culture using the RAPD technique, and no significant result has been found compared to the control. Using RAPD and CREDRA techniques, the genotoxic effect of dicamba has been examined in wheat (Triticum aestivum L.) . Authors have reported DNA alterations in this species. Studies on the genotoxic effect of dicamba by the comet test are extremely limited. In one of these studies, Reynoso et al.  have investigated the genotoxic activity of dicamba-atrazine mixture by comet assay on Zea mays L. and have confirmed significant induction of genetic damages in corn plants at all the concentrations used over the negative control. The genotoxic potential of Dicamba and 2.4 D have been investigated using RAPD and Comet assays and found that both herbicides generated similar damages in the genomic DNA of beans . Çakmak et al.  examined the DNA damaging potential of dicamba in callus obtained from anther culture of sunflower (Helianthus annuus L.). They have revealed the genotoxic effect of dicamba by the comet test.
Due to there is no available literature on A. cepa evaluating the genotoxic potential of Picloram and Dicamba under in vitro conditions, this experiment aimed to determine the genotoxic effect of different concentrations of Dicamba and Picloram used in tissue culture and germinated in distilled water on A. cepa using the Comet test. It is understood that the results would give important insights into the use of dicamba and picloram in both tissue culture and agricultural practices.