Effective Combinations of Plant Growth Regulators for Somatic Embryogenesis from Spinach Root Segments

Abstract

To establish a plant-regenerating system through frequent somatic embryogenesis, a detailed investigation was undertaken on the effects of kinds, concentrations, and combinations of growth regulators on callus and somatic embryo formation from spinach root segments. Although no somatic embryogenesis occurred on the culture medium containing auxins alone, a low frequency of somatic embryo formation was shown on culture media containing auxins combined with natural cytokinins, namely 10 ,ƒÊM NAA with 10 ƒÊM zeatin, 30 ,ƒÊM NAA with 10 ,ƒÊM 2-iP, and 1 ƒÊM 2, 4-0 with 0.1 ƒÊM zeatin or 10 ƒÊM 2-iP. Somatic embryos also formed on culture madia containing auxins combined with GA3. From 50 to 70% of the root segment differentiated somatic embryos on culture media containing 10 to 30 ƒÊM NAA combined with 1 to 100 ƒÊM GA3. These results show that spinach somatic embryogenesis is most facilitated by NAA combined with GA3.

Optimization of micropropagation and <i>Agrobacterium</i>-mediated gene transformation to spinach (<i>Spinacia oleracea</i> L.)

“…So our cultivar that improved in seed and plant improvement institute has a suitable regeneration capacity and we can use this cultivar for transformation program. In addition reports on spinach regeneration through somatic embryogenesis and organogenesis previously indicated that the type of explants plays an important role so root segments are superior in somatic embryogenesis while cotyledons are efficient in organogenesis [6,7,9,11]. However in our study root explants was superior but without transformed explants so we propose the using cotyledon explants which show the higher percent of callus with blue spot and number of blue spot per explants (Figure 1) as transformation indicator ( Table 2).…”

“…Papers dealing with the in vitro regeneration of spinach indicated that genotype is one of the most important factors affecting the regeneration capacity of spinach [4][5][6][7][8][9][10]. So our cultivar that improved in seed and plant improvement institute has a suitable regeneration capacity and we can use this cultivar for transformation program.…”

“…Many papers dealing with the in vitro regeneration of spinach appeared, and it became increasingly clear that genotype is one of the most important factors affecting the regeneration capacity of spinach [4][5][6][7][8][9][10]. However, in addition to the high variability observed in the regeneration capacity of different spinach cultivars, high individual variability within a population of the same spinach cultivar has been reported [10].…”

“…Reports on spinach regeneration through somatic embryogenesis and organogenesis have indicated that the type of explants plays an important role so root segments are superior in somatic embryogenesis while cotyledons are efficient in organogenesis [6,7,9,11]. Leaf discs, hypocotyls, root segments, leaf-derived protoplasts, thin cell layers from hypocotyls and roots have also been utilized in organogenesis [4,5,8,9,[11][12][13][14].…”

The combined effect of photoperiod, light intensity and GA3 on adventitious shoot regeneration from cotyledons of spinach (Spinacia oleracea L.)

“…Regenerated plants obtained by somatic embryogenesis of explants transformed by root inducing transfer-DNA (Ri T-DNA) of Agrobacterium rhizogenes showed abnormal phenotypes (Ishizaki et al 2002). In spinach regeneration, the role of GA 3 has been highlighted in most occasions in both somatic embryogenesis and organogenesis (Al-Khayri et al 1992;Komai et al 1996b;Molvig and Rose 1994;Xiao and Branchard 1993). The effect of exogenous ethylene on embryogenic callus formation in the presence of GA 3 has also been interpreted to facilitate cell response to GA 3 (Ishizaki et al 2000).…”