Somatic Plantlets Research Articles

Somatic embryogenesis from immature and mature zygotic embryos and from cotyledons and needles of somatic plantlets of Larix

Induction of embryonal masses was achieved from full-sib immature zygotic embryos of Larixdecidua Mill., Larix × eurolepis A. Henry, and Larix × leptoeuropaea, and it was found to be affected by the developmental stage of the embryos. Furthermore, the expiant response depended on a specific combination of parental trees used for production of seeds. For the first time, induction was successful in L. decidua with mature zygotic embryos. These embryos were isolated from one of the three seedlots tested and induction frequency was 5%. Mature somatic embryos of L. × leptoeuropaea expressed an initiation frequency of 83%. Cotyledons and needles of plantlets derived from somatic embryos of L. × leptoeuropaea were used as expiants to induce somatic embryogenesis at a frequency of 8 and 3%, respectively. A cytokinin pretreatment seemed to enhance the frequency of induction. Mature somatic embryos were obtained from embryonal masses derived from expiants of different ontogenic age. Future opportunities of research into the induction of embryonal masses from mature tree tissues are discussed.

An improved method for somatic plantlet production in hybrid larch (Larix � leptoeuropaea): Part 2. Control of germination and plantlet development

Germination and plantlet development in somatic embryos of Larix x leptoeuropaea were affected by the duration of the maturation treatment and the concentrations of sucrose and abscisic acid in the maturation media. Extension of the maturation period from 3 weeks to 4 weeks resulted in a significant decrease in germination and plantlet development frequencies. There was no significant effect of abscisic acid concentration on either the number of somatic embryos germinated or the number of plantlets obtained, but it affected the rapidity of the epicotyl development. Sucrose at 0.2 M, applied during maturation, was significantly more beneficial in attaining high germination rates than at 0.1 M. High germination rates (92 and 93%) and plantlet development rates (74 and 80%) were achieved when somatic embryos were matured for a 3-week period on media with either 40 or 60 μM abscisic acid, respectively, and 0.2 M sucrose prior to transfer to the growth regulator-free germination medium. Two acclimatization methods were applied: the first required 10 to 12 weeks and ensured 97% plantlet survival under greenhouse conditions; the second required 2–3 weeks and ensured 86% plantlet survival. This represents the first detailed study of the effects of maturation regimes on the recovery of somatic embryo-derived plants of Larix.

Influence of growth regulators on somatic embryogenesis, plantlet regeneration, and post-transplant survival of Echinochloa frumentacea

After placement on Murashige and Skoog's basal medium supplemented with 3-5 mg/l 2,4-D, immature inflorescence expiants of Echinochloa frumentacea gave rise to three distinct types of callus: a) loosely arranged and soft; b) compact and translucent; c) compact, sticky and mucilaginous. Somatic embryo formation occurred in type 'b' callus in about 18-24 d. Callus types 'a' and 'c' did not produce somatic embryos. The highest percentage of cultures exhibiting somatic embryogenesis occurred on the medium containing 5 mg/l 2,4-D and 0.5 mg/l kinetin. Somatic embryos also formed directly on the inflorescence (without intervening callus formation) in about 15% of the expiants placed on this medium. The addition of paclobutrazol or uniconazole (0.25 or 1 mg/l) to the medium had no influence on the percentage of cultures exhibiting direct somatic embryogenesis, but paclobutrazol slightly increased the mean number of somatic embryos per culture. Many of the callus-derived somatic embryos germinated when subcultured on basal MS medium supplemented with kinetin. Addition of paclobutrazol or uniconazole to the culture medium at 0.25 or 1 mg/l decreased somatic embryo germination and shoot elongation but increased root length and leaf width. Both paclobutrazol and uniconazole increased survival of the plantlets following transplanting to soil. Increased post-transplant survival was accompanied by reduced water loss from plantlets produced on culture media containing triazoles.

Somatic embryogenesis and plant regeneration from zygotic embryo culture in blue spruce (Picea pungens Engelman.)

Somatic embryogenesis and plantlet formation have been achieved from cultured mature zygotic embryos of blue spruce (Picea pungens Engelman.). The effect of three basal media LP, LM, and BLG, all used at half-strength, was tested at the induction phase. LM medium induced somatic embryogenesis to a higher extent than LP whereas BLG did not produce any embryonal-suspensor mass representing stage 1 somatic embryos. The embryonal-suspensor mass was induced on a wide range of auxin/cytokinin ratios. However, media containing either 2 μM NAA and 10 μM BA, or 10 μM NAA and 5 μM BA produced somatic embryos that gave the highest frequency of plantlets. The level of ABA required in the maturation medium for somatic embryos to mature properly varied with the auxin/cytokinin levels in the induction medium on which the somatic embryos were derived. Inclusion of AgNO3 (10 - 100 μM) in the induction medium reduced somatic embryogenesis and embryo conversion.

In VitroSomatic Embryogenesis and Plant Regeneration inClitoria ternatea

A sustainable plant regeneration system in vitro through somatic embryos from mature sexual embryos has been reported in Clitoria tematea. Somatic embryos developed through callus from seedling roots on hormone-free MS medium (MS,). Addition of growth hormones, KN 0-5 mg dm-3 (MS2) or K.N + 1AA 0-5 mg dm^3 of each (MS3) induced direct somatic embryos, in high frequency, on split root and hypocotyl systems. The embryogenie potential varied with the organ, roots or hypocotyls, and also with the medium. The morphogenetic capacity of the somatic embryos is retained for more than 2 years by subculturing at intervals of 4 weeks on MS3 in complete darkness. Somatic embryos, under the appropriate subculture conditions (16 h light/8 h dark photoperiod at 24+ 1 °C on media MS3, MS4 and MS5), resulted in recurrent-somatic embryogenesis and was profuse at the shoot and root apices of the somatic embryos. Mature somatic embryos were transplanted to MS, to stimulate germination and plantlet regeneration. Plantlets, developed from primary and secondary embryos on MS,, were successfully hardened and grown in natural outdoor conditions. The morphology and histology of the somatic embryo and plantlet and the culture conditions for continuous production of plantlets through direct somatic embryogeny are discussed.

Soybean somatic embryo maturation: composition, respiration and water relations

AbstractEmbryogenesis was induced on cotyledons of immature zygotic embryos of soybean (Glycine max(L.) Merrill) placed on solid medium containing 62.5 mmglutamine, soybean seed growth medium salts and vitamins, and 40 mg I−12,4-dichlorophenoxyacetic acid (2,4-D) plus 175 mmmaltose, or 8 mg I−1α-naphthaleneacetic acid (NAA) plus 88 mM sucrose. Somatic embryo development was continued in liquid medium containing 0.16 mg I−1indole-3-butyric acid and 2.64 mg I−1abscisic acid, glutamine and salts as above, and 88–438 mM sucrose in progressively increasing steps. Germination was on solid half-strength Murashige-Skoog medium. During maturation, somatic embryos mimicked zygotic embryos in colour, protein concentration, water and solute potentials, and respiration. Protein and lipid accumulated to 329 and 86 g kg−1dry weight in somatic embryos. Fatty acid composition was similar to that of axes of mature seeds. Before desiccation, the water and solute potentials of maturing somatic embryos declined to −1.13 and −1.99 MPa while turgor increased to 0.86 MPa. Concomitantly, a 60% reduction in activity of the cytochrome oxidase pathway of respiration occurred with somatic embryo maturation at 600 g water kg−1fresh weight. Although small (about 8 mg per embryo), 60% of the somatic embryos formed roots and shoots during germination after maturation without drying and 30% germinated after drying to 60 g water kg−1fresh weight. In the greenhouse, somatic plantlets grew to mature plants with seeds.

Induction of embryogenesis in callus from immature embryos of Persea americana

Both the external morphology and internal anatomy of developing somatic embryoids and plantlets of Persea americana (avocados) were studied. The embryogenic callus, which is embryoidal in origin, was made up of normal parenchymatous tissue toward the centre of a clump of cells and numerous proembryoidal cells on the periphery of the callus mass. Embryogenesis appeared to begin in single cells on the periphery of the callus and from cells on the surface of existing proembryoids. Embryo initials could be recognised by their conspicuous starch contents and by their thickened walls, which lack plasmodesmata. Cell divisions occurred in cells with thickened walls to give rise to globular proembryoids. Proembryoids developed into heart-shaped, torpedo-shaped and cotyledonary embryoids and finally into complete plantlets.

Isolation and Culture of Protoplasts of Panicum maximum Jacq. (Guinea Grass): Somatic Embryogenesis and Plantlet Formation

Summary Protoplasts were isolated from predominantly embryogenic cell suspension cultures derived from proliferating immature embryos and young inflorescences of Panicum maximum Jacq. (Guinea Grass). A large number of protoplasts divided in a liquid culture medium to form hundreds of embryogenic cell aggregates and many globular embryoids. Upon transfer to an agar medium an embryogenic callus was obtained which differentiated more organized embryoids. These embryoids subsequently germinated precociously to form green plantlets. The importance of using embryogenic cell suspensions as a source of totipotent protoplasts of cereals and grasses is emphasized.

Somatic Embryos and Plantlets from an Ancient Clone of the Grapevine (cv. Cabernet-Sauvignon) by Apomixisin vitro

Somatic Embryos and Plantlets from an Ancient Clone of the Grapevine (cv. Cabernet-Sauvignon) by Apomixis<i>in vitro</i>