Suspension cultures which maintained embryogenic potency for more than 18 months were established from excised immature embryos of rice (Oryza sativa L. cv. Konansou). The cultures were subcultured every three days in N6 medium supplemented with proline (10 mM), casein hydrolysate (300 mg/l), sucrose (30 g/l) and 2,4-D (1 mg/l). The frequency of embryogenesis from the embryogenetic suspension cultures reached about 90% when cell clusters (about 1 mm in diameter) were transferred to a solid medium which consisted of N6 medium, NAA (1 mg/l), kinetin (5 mg/l), sucrose (30 g/l) and Gelrite (2 g/l). When smaller clusters of cells (approximately 200–400 μm in diameter) were transferred to a liquid medium which consisted of salts of N6 medium diluted with an equal volume of water plus sucrose (45 g/l), NAA (0.01 mg/l) and 4-PU (0.1 mg/l) at a cell density of 13 clusters/ml in 2 ml of medium, somatic embryogenesis was initated at high frequency (about 50%). Morphological evidence is provided to demonstrate that the regeneration occurred via embryogenesis. This is the first report of high-frequency embryogenesis in suspension cultures of rice cells.

Establishment of a system of high-frequency embryogenesis from long-term cell suspension cultures of rice (Oryza sativa L.).

The seed is the primary organ for the perpetuation of germplasm and propagation in all flowering plants. It contains the embryo which develops from the zygote following the fertilization of the egg cell by one of the male gametes. Gene recombination during the formation, as well as the fusion, of the gametes plays a critical role in the evolution of the angiosperms. During its development the embryo is nourished by the endosperm, which itself is formed as a result of the fusion of the second male gamete with two nuclei (in most of the angiosperms) of the central cell of the embryo sac. During the course of evolution many species have developed special means of propagation to overcome physical, environmental and genetic factors that prevent flowering and seed formation. These include the formation of perennial structures such as tubers, rhizomes, bulbs, stolons, etc (see Sharma and Thorpe, this volume). An advanced form of perennation is found in many apomictic species which produce embryos from unfertilized gametic or somatic cells. Like the zygotic embryos, the apomictic embryos develop within the seed and are protected and dispersed like natural seed. Apomictic embryos of all genotypes — irrespective of heterozygosity — developing from unreduced and unfertilized somatic or gametic cells (agamospermy, adventive embryony, apospory, diplospory) breed true and give rise to clonal populations. Apomixis is known to be under genetic control, but environmental conditions such as temperature and photoperiod may also promote a switch from sexual to apomictic development [1,2].

https://link.springer.com/content/pdf/10.1007/978-94-011-0485-2_10

Somatic Embryogenesis in Herbaceous Dicots

Efficient Plant Regeneration by Somatic Embryogenesis from Root Callus Tissues of Rice (Oryza sativa L.)

It has been known for over 25 years now that plant cell and tissue cultures undergo genetic erosions and show changes of various types, especially in chromosome numbers and ploidy level (Partanen 1963; D’Amato 1965; Murashige and Nakano 1966). However, till the mid 1970’s such changes were considered undesirable and were therefore discarded because the main emphasis was on clonal propagation and genetic stability of the cell cultures. Extensive studies conducted during the last decade have shown that the cell cultures, especially on periodical subculturing, undergo various morphological and genetic changes, i.e., polyploidy, aneuploidy, chromosome breakage, deletions, translocations, gene amplification, inversions, mutations, etc. (see Nagl 1972; Meins 1983; D’Amato 1985). In addition, there are changes at the molecular and biochemical levels (Day and Ellis 1984; Landsman and Uhrig 1985; Ball and Seilleur 1986) including changes in the DNA (Berlyn 1982; Cullis 1983), enzymes (Brettell et al. 1986b), gliadin (Cooper et al. 1986), etc. Since the publication of Larkin and Scowcroft (1981) there has been an upsurge of interest, and the attitude toward these changes has shifted to using the somaclonal variations for plant improvement; they are intentionally induced and much sought after.

Somaclonal Variation — Origin, Induction, Cryopreservation, and Implications in Plant Breeding

Factors affecting tissue culture initiation from maize tassels

An embryogenic callus was obtained from immature panicle of an interspecific hybrid (Oryza sativa x O. latifolia) F1. The medium consisted of HE salts supplemented with 2,4-D, NAA (each 2 mg/l), kinetin (3 mg/l), yeast extract (1360 mg/l) and casein hydrolyzate (300 mg/l). The callus was milk-white in colour compact and granulate in texture. Various developmental stage of embryoid, such as globular, heart-shape, scutellum-shape and mature embryoid were observed in an embryogenic callus. Plantlets were successfully regenerated from 1-month-old callus with more than 80% regenerational frequency in each subculture for 12 passages.

Somatic embryogenesis and plant regeneration in an interspecific hybrid of Oryza.

Cultured immature panicles of rice formed plantlets from spikelets without callus or embryoid formation on MS and HE media containing 2 mg/l each of NAA and kinetin. Developmental stage, ploidy of explant and plant growth regulators in the medium are the major factors affecting the frequency of spikelet budding in young panicle culture. It is suggested that spikelet budding occurs by the reversion of floral primordia to vegetative stage or by the formation of adventitious buds from epidermal cells.

Direct development of plantlets from immature panicles of rice in vitro.

Using MS medium supplemented with 6% sucrose and hormones, plantlets were regenerated from the expiants of mature seeds and young panicles of IRs and IR54. Out of 157 regenerated plants (R1), three were found to be male sterile (ms): one from IRs and two from IR54, including a fertile and sterile chimaera. In the second generations (R2) of IR24 and IR54, one line from each segregated into male sterile and fertile plants. These ms plants could be divided into two types with pollen failure: pollen free (without pollen) and pollen abortive. IR24 was a semi-restorer for ms-plants of the pollen free type derived from the second generation of IR54 somaclones. The segregation ratio of fertile: sterile in both R2 of line 91 and the F2 of ms-plant/IR24 fitted the formula 15/16∶1/16 quite well, showing that the male-sterile was controlled by two independent nuclear genes. Until now, as we know, male-sterile could be produced by hybridization or mutagenesis: sometimes it could be found in nature by spontaneous mutation. Recently the cytoplasmic male-sterile of tobacco was produced by protoplast fusion. This is the first paper to report male-steriles in regenerated plants and their offspring obtained from somatic cell culture.

W. Y. Chen

Papers

Somatic embryogenesis and plant regeneration in an interspecific hybrid of Oryza.

Direct development of plantlets from immature panicles of rice in vitro.

Male sterile mutant from somatic cell culture of rice