تاثیر نوع و غلظت‌های مختلف تنظیم‌کننده‌های رشد گیاهی بر میزان تولید کالوس گیاه رازیانه (foeniculum vulgare) و ترکیبات بیوشیمیایی و متابولیت‌های ثانویه در شرایط درون-شیشه‌ای

گیاه رازیانه (l. foeniculum vulgare) عضوی از خانواده apiaceae است که به علّت داشتن ترکیبات شیمیایی و دارویی شناخته می‌شود. در این پژوهش ابتدا شرایط بهینه جهت تولید کالوس از ریزنمونه‌‌های حاصل از شاخساره گیاه رازیانه مورد بررسی قرار گرفته و سپس ترکیبات بیوشیمیایی کالوس‌های حاصل اندازه‌گیری شد. جهت تولید کالوس ریزنمونه‌‌های حاصل از شاخساره گیاه رازیانه روی محیط کشت ms حاوی تنظیم‌‌کننده‌‌های رشد گیاهی مختلف kin یا bap (صفر، 0.1 و 0.5 میلی گرم بر لیتر) به تنهایی و یا در ترکیب با 2,4-d یا naa و یا iba (هر یک در غلظت‌های صفر، 0.5، 1، 2 و 4 میلی گرم در لیتر) کشت شدند و درصد تولید کالوس، ریشه و اندام هوایی و ترکیبات بیوشیمیایی نمونه کالوس های حاصل (میزان فعالیت آنزیم های پراکسیداز و کاتالاز، میزان تجمع اسید آمینه پرولین، فلاونوئید کل و آنتوسیانین) پس از 3 الی 4 هفته ثبت شدند. طبق نتایج به دست آمده در این پژوهش مشخص شد که بیشترین درصد تولید کالوس (100 درصد) مربوط به محیط کشت حاوی kin (غلظت های 0.1 یا 0.5 میلی گرم بر لیتر) در کنار naa (غلظت های 1 یا 2 یا 4 میلی گرم بر لیتر) بود. از نظر درصد تولید ریشه از کالوس نیز بهترین (100 درصد) مربوط به محیط کشت ms+ 2,4-d (2mg/l)+ bap (0.1mg/l) بود. از سوی دیگر مشخص شد بیشترین میزان فعالیت آنزیم پراکسیداز و کاتالاز و اسیدآمینه پرولین مربوط به محیط کشت ms+ iba (1mg/l)+ bap (0.5mg/l) است. همچنین بیشترین مقدار فلاونوئید در محیط کشت ms+ naa (2mg/l)+ bap (0.5mg/l)  مشاهده شد. طبق نتایج حاصل، استفاده از تنظیم کننده های رشدگیاهی تاثیر مثبت و معنی داری بر مقدار تولید کالوس، تولید ریشه و اندام هوایی و نیز ویژگی های بیوشیمیایی گیاه رازیانه داشت. در پژوهش حاضر، بیشترین میزان ترکیبات بیوشیمیایی به ویژه میزان فلاونوئیدکل و محتوای آنتوسیانینی در کشت درون ‌‌شیشه‌‌ای گیاه رازیانه با افزایش غلظت اکسین و سیتوکنین همراه بود.

the effect of different types and concentrations of plant growth regulators on the amount of callus tissue production by fennel (foeniculum vulgare) explants in vitro

the fennel plant (foeniculum vulgare l.) is a member of the apiaceae umbrella family. in this research, the effect of different plant growth regulators on the production of callus tissue from fennel explants was investigated and then the effect of selected plant growth regulators on the biochemical compounds and secondary metabolites of the resulting callus tissue was evaluated. the desired explants after transfer to the laboratory and surface disinfection on the ms base culture medium containing kin and bap in concentrations (zero, 0.1, and 0.5 mg/l) and 2,4-d, naa, and iba (at concentrations of zero, 0.5, 1, 2 and 4 mg/l) were cultivated. according to the results, the use of kin instead of bap (in lower concentrations) and naa (in higher concentrations) increased the percentage of callus production. according to the results, the highest percentage of callus production (100%) is related to the culture medium containing kin (in concentrations of 0.1 or 0.5 mg/l) along with naa (in concentrations of 1, 2, or 4 mg/l). in this research, it was found that the use of auxin and cytokinin (in combination with each other) increased the induction and production of callus tissue. also, in terms of the activity level of peroxidase and catalase enzymes, and accumulation of amino acid proline, flavonoid, and anthocyanin, a significant difference was observed between different plant growth regulators. so the highest peroxidase and catalase enzyme activities and proline amino acid accumulation are related to ms+1(mg/l) iba +0.5(mg/l) bap culture medium. also, the highest amount of flavonoid was related to ms+2(mg/l)naa+0.5(mg/l)bap culture medium. the highest amount of anthocyanin was related to ms+2 (mg/l)2,4-d+0.5(mg/l)bap culture medium.introductionfennel (foeniculum vulgare l.) is a member of the apiaceae family, which is known for having significant contents of phytochemicals, including flavonoids, coumarins, quinones, phenolic and alkaloid compounds. plant tissue culture is the in vitro culture of cells, tissues, organs, or the whole plant under sterile conditions on a nutrient culture medium of known composition, which is often used to produce plant clones. the resulting clones are genetically uniform and very similar to the mother plant. callus culture is the most common type of culture in plant tissue culture. callus is a growing mass of undifferentiated and unorganized cells, which are formed because of a wound on the surface of the plant. callus tissue can be obtained from all the organs of a whole plant such as meristem, stem tip, lateral buds, leaf, leaf, etc. several factors including genetic structure, type of culture medium, environmental factors (light, ph, and temperature), and plant growth regulators are effective on tissue culture and micropropagation of plants. plant growth regulators, which are used in very low concentrations, are effective in the distribution of substances inside the plant, cell division, and cell growth. plant growth regulators, especially auxins and cytokinins, play a very important role in controlling the growth and development of cells both in the whole plant and in explants cultured in vitro. material and methodsin this research, first, the optimal conditions to produce callus from explants obtained from fennel shoots were investigated, and then the biochemical compositions of the resulting callus were measured. to produce callus tissue, explants from f. vulgare shoots were prepared from the medicinal plant research center of ardabil university of medical sciences. the samples were first examined phenotypically and after selecting fresh samples without obvious contamination, they were washed with a dishwashing solution for 30 minutes and then rinsed under running water for one hour. the explants were then immersed in h2o2 solution (5%) for 10 minutes and after rinsing twice with sterile water (autoclaved twice) for 10 minutes, they were disinfected in 2.5% sodium hypochlorite. at the end, sterilized explants on ms culture medium containing different plant growth regulators kin or bap (zero, 0.1 and 0.5 mg/l) alone or in combination with 2,4-d or naa or iba (each in concentrations of zero, 0.5, 1, 2 and 4 mg/l) were cultivated and the percentage of callus production, root and shoot and the biochemical composition of the resulting callus samples (peroxidase and catalase activity levels), the amount of amino acid accumulation of proline, total flavonoid and anthocyanin) were recorded after 3-4 weeks. this research was conducted as a completely randomized design experiment with three replications, and the data analysis was done using spss ver. 26.results and discussionaccording to the results obtained in this research, it was found that the percentage of callus production from explants obtained from fennel plant shoots, roots, and aerial parts obtained from callus, as well as the weight of the collected callus, had a significant difference between most of the plant growth regulators treatments and the control treatment. according to the results obtained in this research, it was found that the highest percentage of callus production (100%) was related to the culture medium containing kin (concentrations of 0.1 or 0.5 mg/l) along with naa (concentrations of 1, 2, or 4 mg/l). according to the results of the study, the use of auxin and cytokinin (in combination with each other) increased the induction and production of callus. in terms of the percentage of root production from callus samples, the best (100%) was related to ms+ 2,4-d (2 mg/l) + bap (0.1 mg/l) culture medium. also, ms culture medium containing 1mg/l of 2,4-d along with 0.5 mg/liter of bap had the highest percentage of shoot production from callus samples of the fennel plant (55.53%). on the other hand, a significant difference was observed in the activities of peroxidase and catalase enzymes, amino acid proline, flavonoid, and anthocyanin accumulation between different plant growth regulators. the highest level of peroxidase, catalase, and amino acid proline was related to ms + iba (1mg/l) + bap (0.5mg/l) culture medium.conclusionaccording to the results, the use of plant growth regulators had a positive and significant effect on the amount of callus production, root, and shoot production, as well as the biochemical characteristics of the fennel plant. in this research, it was found that the use of kin and naa causes the maximum increase in callus production from fennel explants. to increase the antioxidant and anticancer properties of callus samples, it is very important to increase the amount of flavonoids and anthocyanins in the callus, and in the current study, the highest amount of biochemical compounds, especially the amount of flavonoids and anthocyanins in the in vitro culture of the fennel plant, was associated with an increase in the concentration of auxin and cytokinin.