In vitro propagation of garlic (Allium sativum L.) using clove segment explants

Introduction

Garlic is the second most widely used cultivated Allium after onion. It has long been recognized worldwide as a valuable spice for foods and a popular remedy for various ailments and physiological disorders. It is used worldwide to flavor various dishes. Allium sativum L., commonly known as garlic, is a member of the onion family (Alliaceae). Garlic has been used throughout the ages for both culinary and medical purposes. Garlic (Allium sativum) is an aromatic monocotyledonous bulbous plant and an annual spice belonging to the subfamily Allioideae of the family Alliaceae. It is used worldwide as a condiment and for its medicinal properties (Mawalkar et al., 2018) [4] . Garlic has an unusually high concentration of sulphur-containing compounds. Sulphur compounds, including allicin (thio2-propene-1-sulfinic acid S-allyl ester), were confirmed to be the main active components in the root bulb of the garlic plant. (Mehta et. al., 2013) [5] It is use in component in the preparation of drug and pesticide. Garlic is an important culinary as well as medicinal herb due to its ability to improve the taste of food and to its antibiotic, antitumor, and antithrombotic effects in animal cells. Garlic (Allium sativum L.) is propagated exclusively vegetatively, a process that has a low coefficient of multiplication and potential for transmission of viral diseases. (Muhammad et al., 1997) Garlic (Allium sativum) is an aromatic monocotyledonous bulbous plant and an annual spice belonging to the subfamily Allioideae of the family Alliaceae. It is used worldwide as a condiment and for its medicinal properties. (Mawalkar et. al., 2018) [4] Garlic is the world's oldest cultivated crop. It is an approximately 5000-year-old indigenous herb originating in Central Asia (Kazakhstan), South Asia, or southwestern Siberia. Garlic is closely related to the onion, shallot, leek, and chive. It is easy to grow and can be cultivated year-round. It is cultivated in both temperate and tropical climates (Mawalkar et al., 2018) [4] . Allicin has a wide range of biological and pharmacological activities, including anticoagulation, antihypertensive, antimicrobial, antibiotic, antiparasitic, antimycotic, antiviral, antitumoral, antioxidant, anti-aging, antiplatelet, detoxification of heavy metals, fibrinolysis, and hypolipidemic (lipid-lowering) activities. It is an immune enhancer and modulator. (Mehta et.al. 2013) [5] Need for propagation of Garlic: The propagation rate of garlic in the field is low; therefore, it takes many years to produce a sufficient number of seed bulbs for the practical cultivation of a new variety. Many garlic cultivars are susceptible to diseases caused by viruses and fungi and suffer from insect pests. In vitro regeneration or micropropagation is the best alternative to overcome these hurdles, and it holds tremendous potential for rapid multiplication and production of high-quality, uniform, and disease-free planting material. (Mawalkar et al. 2018) [4] Materials and Methods Source of explant and surface sterilization: Clove segments were collected as explants from mature, healthy cloves from DOGR, Pune, and brought to the tissue culture laboratory of the Department of Plant Biotechnology, College of Agricultural Biotechnology, Loni. These explants were thoroughly washed under running tap water 4-5 times. They were then treated with 0.1% Bavistin for 3 min. In the next step, the samples were washed with Tween 20 for 2-3 min. Surface sterilization was performed with mercuric chloride (HgCl 2 ) (0.1%) for 30 s under aseptic conditions and finally washed 3-4 times with sterile distilled water to remove traces of HgCl 2 .and these sterilized explants were cultured on MS medium with different concentrations of growth regulators.

Initiation stage and culture conditions:

Clove segments in the initiation stage were transferred to four different MS media supplemented with BAP (0.5-2.0 mg/l) in combination with Kn (0.5 mg/l). The inoculated culture bottles were incubated in an incubation room with sterile controlled environmental conditions, such as temperature 25±2 °C, relative humidity (RH) 70%, 16 h photoperiod/day with 2000-3000 lux light intensity under cool white fluorescent tubes (Philips, India).

Shoot elongation:

Different hormones, such as BAP and Kn, with multiple combinations, were successfully used for Shoot Elongation. Shoots were transferred to fresh MS medium with different concentrations of BAP and Kn for elongation.

Rooting:

The explants were transferred for rooting in MS media supplemented with two different concentrations of IBA.

Results and Discussion

Shoot initiation: Data obtained after 7 days of culture initiation showed that shoots of Allium sativum could be established on all tested media, including the medium. The best result was obtained on MS medium supplemented with 2.0 mg/l BAP + 0.5 mg/l Kn. (Table 1). The shoot length (average shoot length was 2.5 cm) was observed on medium supplemented with 2.0 mg/l BAP + 0.5 mg/l Kn (Table 1,

References

1. Bekheet SA. Micropropagation of garlic (Allium sativum). Ann Agric Sci. 2004;49(1):223-232.
2. Hassan MN, Haque MS, Hassan MM. An efficient protocol for somatic embryogenesis of garlic (Allium sativum L.) using root tip as explant. J Bangladesh Agric Univ. 2014;12(1):1-6.
3. Khan N, Alam MS, Nath UK. In vitro regeneration of garlic through callus culture. J Biol Sci. 2004;4(2):189191.
4. Mawalkar M, Chavan NS, Jagtap RR, Kharade SS. Rapid in vitro plant regeneration of Allium sativum. J Pharmacogn Phytochem. 2018;7:3087-3089.
5. Mehta J, Sharma A, Sharma N, Megwal S, Sharma G, Gehlot P, et al. An improved method for callus culture and in vitro propagation of garlic (Allium sativum L.). Int J Pure Appl Biosci. 2013;1(1):1-6.
6. Metwally EI, El-Denary ME, Dewir YH, Naidoo Y. In vitro propagation of garlic (Allium sativum L.) through adventitious shoot organogenesis. Afr J Biotechnol. 2014;13(38):3892-3900.
7. Haque MS, Wada T, Hattori K. High frequency shoot regeneration and plantlet formation from root tip of garlic. Plant Cell Tissue Organ Cult. 1997;50(2):83-89.
8. Seabrook JE. In vitro propagation and bulb formation of garlic. Can J Plant Sci. 1994;74(1):155-158.