In Vitro Micropropagation Protocol Optimization of Garlic (Al lium sativum.L) ?�?Holetta Local?�? and ?�?Kuriftu?�? Varieties Using Shoot Tip Culture

Abstract

Garlic (Allium sativum L.) is major bulb plant predominantly used for flavoring and serving dishes and also used as a medicinal plant all over the world. However, numerous crop management problems, the nature of propagation, diseases and insect pest limited its production. Virus is a major detriment for reduction of garlic yield and quality. To produce virus-free garlic, shoot tip culture has been widely adopted. However, shoot tip culture require the optimized protocol and necessary cultural conditions. The objective of this study was to optimize an efficient protocol for in vitro micropropagation of selected garlic varieties (‘Holeta Local’ and ‘Kuriftu’) through direct organogenesis using shoot tip culture. The garlic cloves were washed with powder soap, sterilized with 70% ethanol and 15% chlorine bleach solution. Shoot tips containing apical meristems 0.5 to 0.8 cm in length were aseptically excised from sprouted cloves and cultured on half strength Murashige and Skoog medium supplemented with 10mg/l Benzyl Amino Purine (BAP) to initiate the growth of shoot tip and shoot induction. For this experiment cytokinin alone was crucial for better direct shoot induction from shoot tip culture. Shoot multiplication was carried out on half strength MS medium containing different concentrations of Benzyl Amino Purine (BAP) in combination with Naphthalene acetic acid (NAA). Half MS medium without PGRs was used as control. Among the treatment, 2.0 mg/l BAP+0.25 mg/l NAA was found to be the best for ‘Holeta Local’ where a maximum of 5.48±0.28 shoots and for ‘Kuriftu’ a maximum of shoot 4.86±0.25 was obtained on 1.5 mg/l BAP+0.25 mg/l NAA. The highest shoot length 7.63±0.73 cm and 6.85±0.33 cm were recorded on 1.5 mg/l BAP+0.25 mg/l NAA for ‘Holeta Local’ and ‘Kuriftu’ respectively. Root initiation was carried out on half strength MS medium containing different concentrations of NAA. Half MS medium without PGRs was used as control. Among the treatment, the highest number of roots 3.42±0.32 and 3.65±0.51 per explant were recorded on 1.0 mg/l NAA for ‘Holeta Local’ and ‘Kuriftu’ respectively. The highest mean length of root per explant 2.08±0.32 cm was obtained on 0.5 mg/l NAA for ‘Holetta Local’ and 2.46±0.53 cm for ‘Kuriftu’ on 1.0 mg/l NAA. The highest percentage of root formation 100% was recorded on 1.0 mg/l NAA for both varieties. Of those shoots which were acclimatized in the greenhouse, 78% of ‘‘Holetta Local’’ and 84 % of ‘Kuriftu’ survived