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Callus induction and proliferation of Alkanna orientalis var. orientalis and Alkanna sieheana containing valuable alkannin/shikonin (A/S) derivates were investigated using leaf base and stem segment explants. Stem segments and cotyledonary leaf base of both species were cultured on Murashige and Skoog medium fortified with different concentrations of BAP, Kn, NAA, IAA and IBA for callus induction and shoot regeneration. High frequency reproducible, prolific and compact calli formation was obtained from the stem segments of both species in all media tested. The frequency variations of callus induction and shoot regeneration were discussed in terms of different species, plant growth regulators and explant resources. A. orientalis and A. sieheana may be considered to be alternative plants for the A/S production in vitro. Introduction The genus Alkanna with 50 species is a member of the Boraginaceae family (Mahmoudi et al. 2012). Turkey is one of the most important gene centres for the genus represented in the flora of Turkey by 41 taxa belonging to 36 species with an 80% endemism rate (Güner et al. 2012). The genus has also herbaceous species and spread out in Mediterranean coast, central, eastern and western of Anatolia. Many species of the genus synthesise red pigments of the isohexenylnaphthazarins which are important plant secondary metabolites. These compounds are commonly known as alkannin/shikonin (A/S) and their derivatives are lipophilic hydroxynaphthoquinone pigments. A/S is enantiomeric hydroxynaphthoquinones and these compounds are found in at least 150 species belonging to the genera Alkanna, Lithospermum, Cynoglossum, Echium, Anchusa and Onosma of the Boraginaceae family (Kumar et al. 2013). Bioactive substances are mainly available in the outer surface of roots. A/S and its derivatives are used for dyeing in areas such as textile, food, cosmetics (Pal et al. 2017). The compounds have also various biological activities such as a strong wound healing (Papageorgiou et al. 2008) antibacterial and antifungal properties (Damianakos et al. 2012), antiinflammatory (Mahmoudi et al. 2012), antioxidant (Wang et al. 2010), as well as antiallergic, antineoplastic, antipyretic and antihydropic activities (Su et al. 2011). Recently it has been known that A/S and its derivatives exhibit antitumor activity (Damianakos et al. 2012) and these compounds are actively used in anticancer treatments in many countries (Yeh et al. 2015). Wild forms of the genus usually contain A/S in their roots in nature. Optimum production of A/S and its derivatives in roots of plant is 7 9 years. Therefore, the production of these compounds with culture is difficult. Also, if the plants were collected from nature, a stable substance can not be obtained because it is collected from different locations and environments. *Author for correspondence: <cennet.yaman@bozok.edu.tr>, <uranbey@ankara.edu.tr>. Department of Field Crops, Faculty of Agriculture, Ankara University, 06100 Ankara/Turkey. Department of Field Crops, Faculty of Agriculture and Natural Sciences, Uşak University, 64200 Uşak/Turkey. Department of Pharmaceutical Botany, Faculty of Pharmacy, Selçuk University 42100 Konya /Turkey.

Callus induction and regeneration of Alkanna orientalis var. Orientalis and A. sieheana