استقرار محیط های کشت سوسپانسیون سلول جنینی سیر (Allium sativum)، تولید مثل گیاهی و تجزیه و تحلیل های بیوشیمیایی Establishment of embryogenic cell suspension cultures of garlic (Allium sativum L.), plant regeneration and biochemical analyses

All commercial garlic (Allium sativum L.) cultivars are sterile (Etoh and Simon 2002; Kamenetsky and Rabinowitch 2001) and must be propagated vegetatively. Consequently, the chance of disease transmission is increased (Novak 1990), particularly as garlic varieties are standardly infected with various viruses (Dolores et al. 2002; Lot et al. 1994). Methods for virus elimination and the production of virus-free material have been developed using meristemtip culture (Ayabe 2001; Chovelon et al. 1990; Senula et al. 2000). Various researchers applying different micro-propagation protocols have obtained a limited multiplication rate with respect to the production of virus-free material (Barandiaran et al. 1999; Haque et al. 1997; Kahane et al. 1992; Kim et al. 2003; Myers and Simon 1998; Nagakubo et al. 1993; Robledo Paz et al. 2000; Zheng et al. 2003). As such, embryogenesis might be applicable in scaling-up the propagation of healthy garlic tissues (Xue et al. 1991); for example, a somatic embryogenesis process was reported recently (Fereol et al. 2002). Nevertheless, to date, the multiplication rates for producing virus-free garlic plants have been too low to satisfy the economic requirements for a practical mass propagation system. Plant production from embryogenic cell suspension culturederived somatic embryos is a possible alternative. Due to the high medium-to-tissue contact in liquid culture system, media effects are rapid, and embryo development can be more strictly controlled than with a solid support system.  There have been limited reports on garlic suspension cultures. Nagasawa and Finer (1988) reported suspension cultures of callus proliferating as nodular clumps, but they did not obtain true cell suspensions or plant regeneration. Barrueto Cid et al. (1994) established garlic suspension cultures and regenerated plants, but they did not mention embryogenic characteristics nor the regeneration frequency. Moreover, their protocol required a high concentration of 2,4-D (1.1 mg/l), thereby risking somaclonal variation (Al Zahim et al. 1999). The objective of the investigation reported here was to establish and characterise a reliable protocol for garlic proliferation via embryogenic cell suspension cultures. We investigated the effect of several parameters on cell multiplication and plant regeneration to determine and optimise conditions for maintaining the cultures. Special attention was directed towards keeping the 2,4-D concentration as low as possible. Histological studies ascertained the embryogenic capacity induced in cell suspension cultures. Chemical analysis was performed on micro bulbs from cell suspension embryo-derived plantlets (CS) and in vitro shoot proliferation-derived plantlets (SP) as a quality trait to distinguish between these two garlic proliferation strategies.