Читать книгу Biotechnology of Fruit and Nut Crops - Группа авторов - Страница 159

The first report of date palm in vitro culture was by Schroeder (1970). Somatic embryogenesis was described by Tisserat (Tisserat, 1979a,b; Tisserat and DeMason, 1980) and Reuveni (Reuveni et al., 1972; Reuveni and Lilien-Kipnis, 1974; Reuveni, 1979). Current protocols have been reviewed (Fki et al., 2011; Mazri and Meziani, 2015; Al-Khayri and Naik, 2017; Gantait et al., 2018).

In early studies, zygotic embryos, immature fruits, roots, leaf petioles, lateral buds, shoot tips, stem pieces and rachilla tissue were used as explants (Tisserat, 1979a; Sharma et al., 1980). The most commonly utilized explants are shoot tips, either of the main trunk or from the offshoots. These are located at the ‘palm heart’ and are responsible for generation of all leaves and inflorescences (Tomlinson, 1990; Cohen, 2017). The harvest of shoot tips requires the sacrifice of the entire palm or the offshoot. An alternative explant source is the immature inflorescence (Bhaskaran and Smith, 1992; Abul-Soad, 2011; Gadalla, 2017). These can be dissected from the tree with only minor and temporary damage, although they are available only for a brief period each year.

Various parameters, including cultivar, the source of explant and the plant growth regulators, affect induction of embryogenic cultures (Gantait et al., 2018). Apical shoot tips, lateral buds and leaf primordia have been used in addition to developing inflorescences. Although several different plant media have been tested, most studies used MS medium supplemented with sucrose as the carbon source. The use of 45–452 μM of synthetic auxin, usually 2,4-dichlorophenoxyacetic acid (2,4-D), alone or in combination with other plant regulators, is essential (Fki et al., 2011; Gantait et al., 2018). Browning of explants is avoided by addition of activated charcoal and PVP into the induction medium. Fine-tuning of plant growth medium and concentrations of carbon source and plant growth regulators is necessary for each cultivar. As a result, many studies have presented optimized protocols for specific cultivars. Darkness is required for embryogenic culture induction and somatic embryo development. Shoot and root development usually occur with a 16 h or 12 h photoperiod with lighting of 30–100 μmol/m²/s. The cultures are maintained at 24–28°C (Gantait et al., 2018).

Although embryogenic cultures are induced on semi-solid medium, they can be maintained as embryogenic suspension cultures in order to maximize recovery of somatic embryo-derived plants (Fki et al., 2003; Zouine et al., 2005; Sané et al., 2006; Zouine and El Hadrami, 2007; Abohatem et al., 2017). Suspension cultures on liquid MS media with reduced level of auxin can generate 2–20 times more somatic embryos in comparison with growth on semi-solid media (Fki et al., 2003; Saker et al., 2007b). The gentle shaking of the culture at c.125 rpm is optimum.

Suspension cultures can be efficient tools to study the effect of abiotic stresses, i.e. salinity and drought. Suspension cultures grown under different concentrations of KCl, NaCl or CaCl₂ or in medium containing polyethylene glycol enabled isolation of embryogenic cultures with enhanced tolerance of salinity and drought (Al-Khayri and Ibraheem, 2014).