Plantlet regeneration via somatic embryogenesis from subcultured callus of mature embryos of Picea abies (Norway spruce (original) (raw)
Related papers
Plant Science, 1985
Embryos ofPicea abies at various developmental stages were cultured on defined media supplemented with 2,4dichlorophenoxyacetic acid (2,4-D) (10 -s M) and N6-benzyladenine (BA) (5 X 10 -6 M). The immature embryos gave rise to a highly friable and embryogenic callus which could be maintained by subculture and contained polarized and organized structures (somatic embryos) consisting of long highly vacuolated cells at one end (suspensor) and a group of small meristematic cells at the other (embryonal end). These structures closely resembled the early stages of normal zygotic embryogeny. Upon further culture these structures formed a bipolar shoot-root axis with an independent and closed vascular system. In many instances either the shoot or the root meristems failed to differentiate. Embryogenic tissues obtained on agar media could be transferred to liquid media and maintained by subculture for at least 6 months. The development of somatic embryos was observed in the liquid cultures also.
Enhancement of somatic embryogenesis in Norway spruce ( Picea abies L.)
TAG Theoretical and Applied Genetics, 1988
Embryogenic callus developed in 55% of the mature embryo explants of Norway spruce (Picea abies L.) growing on a LP medium minus the amino acids and sugars (except sucrose). This is the highest reported yield of embryogenic callus from mature embryos of P. abies that has ever been reported. Callus induction from either the middle or the end of the hypocotyl of the embryos began after 2-3 weeks. Three types of calli were recovered: (a) globular, (b) light green-compact, (c) white mucilaginous. Only the white mucilaginous calli were embryogenic. The globular and light green-compact calli never become embryogenic, even after several subcultures. The development of somatic embryos was accomplished on half-strength macro-elements of NSIII medium containing 1 gM e-naphthaleneacetic acid, 1 gM abscisic acid, and 3% sucrose. The addition of 10-7M buthionine sulfoximine to the medium increased the development of somatic embryos by three fold. These results suggest that there is a great potential for increasing the frequency and development of somatic embryos in P. abies. Careful selection of the genotype and modification of the culture medium is required.
1987
of white spruce (Picea glauca) somatic embryos from protoplasts derived from an embryogenic suspension culture was accomplished using a c~lture medium containing 2 mgl -~ 2,4-D and I mgl-6-BAP. Divisions within 2 days led to plating efficiencies in the order of 24% after 9 days. A reduction in the osmoticum, necessary for sustained growth, was carried out gradually over 30 days. Embedding in agarose and culture in 5 cm petri dishes prior to transfer of agarose blocks to a bead type culture, led to the formation of somatic embryos as early as 23 days after isolation and yielded plating efficiencies in the order of 5-10% after 35 days culture.
Somatic embryo maturation from long-term suspension cultures of white spruce (Picea glauca)
In Vitro Cellular & Developmental Biology - Plant, 1993
The production of cotyledonary somatic embryos of white spruce from cultures grown long-term as suspensions was investigated. We report the effects of removal of 2,4-dichlorophenoxyacetic acid (2,4-D) from the maintenance medium (ordinarily containing both 2,4-D and benzyl adenine) before (+_)-ABA-stimulated maturation. In particular the use of a 1-wk culture period without 2,4-D was found to improve the production of normal-looking cotyledonary somatic embryos. Using high performance liquid chromatography analyses of culture supernatants, it was determined that this affect was not related to altered ABA metabolism. Germination of cotyledonary somatic embryos from cultures pretreated by the 1-wk culture period without 2,4-D was improved compared with similar embryos from cultures that had not been pretreated.
Tissue culture techniques enable mass propagation of elite cultivars of date palm (Phoenix dactylifera L.). The main limitations for date palm in vitro multiplication are the low rates achieved using solidified medium and the long period needed to produce acclimatized plantlets. This research focuses on the comparison between different culture types and plant growth regulator (PGR) combinations on callus growth and somatic embryo formation of cv. Zaghlool. For callus growth, 200 mg friable embryogenic callus dispensed in Rasotherm and Phytacon flasks containing 200 ml liquid (100 rotations per minute) and in temporary immersion system, RITA® bioreactor (5 min immersion every 12 h), were compared with cultures grown on 200 ml solidified medium. For somatic embryo formation, 500 mg of friable embryogenic callus grown in Erlenmeyer flasks filled with 50 ml liquid or solid MS medium. The medium was supplemented with 0.1 mg l-1 Naphthaleneacetic acid (NAA), 1.5 g l-1 activated charcoal (AC) with or without 0.05 mg l-1 6-Benzyl amino purine (BAP) compared to PGR-free medium. Results proved that cell suspension cultures produced the highest callus fresh mass as compared to the other systems tested, and the callus fresh mass reached 4 g after 16 weeks. The temporary immersion system did not significantly enhance the fresh mass of callus compared to the solidified medium. For somatic embryo induction, the number of somatic embryos increased in cell suspensions 6-16 fold compared to the solidified medium. Using liquid MS medium enriched with 0.1 mg l-1 NAA and 1.5 g l-1 AC gave rise to the highest number of somatic embryos formed from 500 mg initial callus: 160 embryos. The number of somatic embryos was also affected by the callus source. The calli induced from leaflet segments excised from converted somatic embryos resulted in a lower somatic embryo number than those of shoot tip origin with about 60 somatic embryos per 500 mg callus. The formation of somatic embryos using liquid medium required only 6 weeks, thus considerably reducing the previously reported period of 18 weeks which is required for somatic embryo formation using solidified media.
Plant Cell, Tissue and Organ Culture, 1992
An elite Chinese cotton (Gossypium hirsutum L.) cultivar Simian-3 was chosen for tissue culture. Callus with a high frequency of somatic embryogenesis, somatic embryos, and regenerative plants was obtained. Callus was induced from three types of explants on MSB (MS salts with B 5 vitamins) medium supplemented with zeatin (ZT) only, but the percentage of callus induction and growth of callus varied. It appeared that it was much easier to induce callus from hypocotyl than cotyledon or root explants. The concentrations of ZT were critical to the induction and proliferation of callus. The optimum ZT concentration for callus induction was 3.0~5.0 mg/L. Two kinds of callus could be identified after 70 days of culture: embryogenic and nonembryogenic callus. Embryogenic callus developed into somatic embryos at various stages after 20 days of subculture. The capability of embryogenesis depended on the explant types. The root was the most responsive explant for production of somatic embryos, the hypocotyl was the next, and the cotyledon was the last. Moreover, a low concentration of ZT was advantageous to the induction of embryogenic callus. 2,4-dichlorophenoxyacetic acid (2,4-D) promoted the proliferation of embryogenic callus, but had a negative effect on the differentiation and germination of somatic embryos. Addition of activated charcoal or a proper combination of ZT and 3-indoleacetic acid (IAA) could promote the production, maturation and germination of somatic embryos. The best medium for the proliferation of embryogenic callus was ZH medium (Zhang et al., 1996) with 1.0 mg/L 2,4-D, 0.5 mg/L kinetin (KT) and 0.5 mg/L ZT. The best medium for the differentiation and germination of somatic embryos was MSB with 0.1 mg/L ZT and 2 g/L activated charcoal. An efficient protocol for the production of high frequency somatic embryogenesis and plant regeneration of an elite cotton variety Simian-3 has been developed. Complete plants could be regenerated through somatic embryogenesis from hypocotyl, cotyledon and root explants in 3-4 months.
Improvement of somatic embryogenesis in highland-papaya cell suspensions
Plant cell, tissue and organ culture, 1996
Axillary buds (2 ram) from 3-year-old Caricapubescens Lenn6 et Koch (highland papaya) fruit-bearing plants grown in the greenhouse were cultivated in N-N-medium supplemented with different growth regulators (naphthaleneacetic acid and indoleacetic acid in combination with Zeatin, benzyladenine, Kinetin and thidiazuron. Several responses were observed within 2-3 months; namely, sprouting of the preformed axillary buds, bud branching into multiple shoots, callus formation at the basal end of the explant and somatic embryogenesis in the preformed callus. Somatic embryogenesis was frequent in most of the tested growth regulator combinations, with the exception of thidiazuron which showed no effect. A much higher yield of somatic embryos could be obtained in suspensions. Somatic embryogenesis was enhanced by the occurence of adventive embryogenesis on single embryos as globular embryo clusters. This was observed in cell suspensions initially grown in a WPM-medium with 2,4-dichlorophenoxyacetic acid, or in combination with benzyladenine or zeatin, for 6 days, then maintained in a growth regulator-free medium under continuous agitation (50 RPM) on an orbital shaker for 3 months. Single cells grown in the absence of 2,4-dichlorophenoxyacetic acid did not initiate embryogenesis and de-differentiated into callus. Plantlets were recovered after transfer of mature embryos from cell suspensions into Magenta flasks. In a second subculture, adventitious embryogenesis occurred spontaneously in clusters at the globular embryo stage under the same growth conditions, yielding a high number of embryos. The culture conditions described above allowed initiation of a large number of somatic embryos directly from cell suspensions through adventive somatic embryogenesis and indirectly from callus on axillary buds.
Somatic embryogenesis and plant regeneration from callus cultures of Cleome rosea Vahl
2010
This paper describes a protocol for the efficient vegetative propagation of Cleome rosea by somatic embryogenesis. Leaf and stem explants from nursery-grown seedlings of C. rosea were cultivated on Murashige and Skoog (MS) medium supplemented with indole-3-acetic acid (IAA), a -naphthaleneacetic acid (NAA), 4-amino-3,5,6-trichloropicolinic acid (picloram) or 2,4-dichlorophenoxyacetic acid (2,4-D). Nodular calli were produced from both explant types in the presence of 4.5 and 9.0 µM 2,4-D. Embryo development and maturation were achieved when calli from stem explants were transferred to media containing a ten-fold reduction of 2,4-D concentration initially used (0.45 and 0.90 µM). Leaf-derived calli did not form embryos with the same treatments. The highest frequency of embryogenic callus formation (85%) and number of embryo per callus (13.45 ± 2.8) were achieved during the first subculture on medium supplemented with 0.90 µM 2,4-D. Embryo conversion into plantlets was achieved follow...
In-vitro developmental biology- Embryo
A simple and efficient protocol has been developed for in vitro regeneration of M. acuminata ssp. burmannica (AA) plants. Somatic embryos were produced when immature and mature zygotic embryo explants were cultured on Murashige and Skoog medium supplemented with plant growth regulators 2,4-dichlorophenoxyacetic acid; (2,4-D), picloram or benzyl adenine and indole acetic acid. In general, immature embryos responded better than mature embryos. Callus proliferation was highest in medium supplemented with 2,4-D (4.5 μM). Subsequent transfer of callus to fresh medium produced rapidly proliferating embryogenic calli. Embryogenic calli were maintained in complete darkness for 15 d followed by cycles of 8 h dark and 16 h light, under white fluorescent lamps with a light intensity of 3,000 lm/m 2 and at temperature of 28±2°C. Regeneration of embryogenic calli into plantlets was higher for immature embryos (76.6%) than for mature embryos (50.6%). This plant regeneration protocol using mature or immature zygotic embryos, via somatic embryogenesis, has significant potential to improve germination efficiencies of hybrid progenies used in conventional breeding strategies. Furthermore, tests on seed storage showed that seed viability rapidly decline after harvesting and was negligible after 9 mo of storage. This indicates using freshly harvested seeds as explant material is necessary for maximizing the tissue culture response.
Somatic embryogenesis from vegetative shoot apices of mature trees of Pinus patula
Embryogenic cultures were initiated and established from apical shoots of mature trees of three genotypes of Pinus patula Scheide et Deppe. Factors affecting initiation, including cold pretreatment, basal medium composition, growth regulators and gelling agent concentration, and the effect of partial desiccation on somatic embryo maturation were investigated. Cold pretreatment of thick sections (0.5–1.0 mm) of api-cal shoots at 2 °C for 3 days on 0.3% activated charcoal induced white mucilaginous embryogenic callus on initiation medium. Subculture of this embryogenic callus on maintenance medium resulted in the formation of embryonal suspensor masses with proembryos. Partial desiccation (12–90 h) of embryo-genic tissue at the proembryo stage of development, prior to transfer to maturation medium containing 9 g l –1 Gellan gum, enhanced somatic embryo maturation and germinability. The frequency of maturation increased from 5.3 to 16.5% after 12 h of desiccation and from 16.5 to 73.8% after 24 h of desiccation, but longer periods of desiccation were ineffective.