C. Bigot

Adventitious shoot production from strips of stem in the Dutch elm hybrid 'Commelin' : Plantlet regeneration and neomycin sensitivity

The regenerative ability of small strips of stem of the Dutch elm hybrid ‘Commelin’ was tested as well as its sensitivity to neomycins. Cambium explants (1 mm thick), were excised from woody stems collected in the field. Up to 20 buds/explant were induced within 2–3 weeks giving 2–5 rootable shoots/explant after 5–6 weeks. Shoot excision every week from week three improved the yield up to 7 shoots per explant. Fourteen and 2.9 μM GA3 promoted shoot growth. Cytokinins (1 μM zeatin or 5 μM BA or 0.05 μM TDZ) completely inhibited shoot production and promoted callus formation. Kanamycin and paromomycin at between 240 and 360 μM inhibited shoot formation as did geneticin at 80 μM. The shoot-forming ability of the explants was high from leaf fall in the autumn to the spring flush, but could be maintained up to September by using cold storage (5–7 °C). Ninety-six percent of the shoots rooted with 0.5 μM IBA and were successfully acclimatized despite having a large basal callus.

Regeneration of Plants from Protoplasts of Ulmus Species (Elms)

Elms belong to the Ulmaceae family. The genus Ulmus comprises about 30 species of usually deciduous trees. They are native to the Northern Hemisphere throughout four continents (Bean 1980). Europe has 5 species, North America 8 species, and no less than 23 species have been identified in Asia where China appears to be the center of diversity of the genus (Heybroek 1981). Natural distribution and habitats of elms range from sea level up to 3900 m and from the edge of the tundra to the tropics in Central America and Southeast Asia.

In vitro budding ability of woody internode and Agrobacterium susceptibility as prerequisites for elm genetic transformation

Summary In order to improve elm resistance to dutch elm disease by genetic engineering, woody internodes of field grown elms Commelin were tested for regeneration capacity and susceptibility to Agrobacterium tumefaciens . After a drastic surface disinfection (especially burning), the explants were introduced in vitro on a medium containing cef otaxime (500 mg - L ∔1 ). Budding took place within 2 weeks on the cambial part of the explants. After 1-2 months, the number of rootable shoots per explant was 3 -6, increasing up to 13 by shoot excision. Shoot rooting (70 %) and acclimatization (survival 80 %) were easy. Agrobacterium tumefaciens wild strains (Ach5, T37, C58, 82-139) were either inoculated on shoots or cocultivated on internodes. Octopine/agropine type Ach5 induced tumors (> 50 %) where agropine was often detected. Efficiency of 82-139 was lower. However, it was very effective in improving budding ability of internodes up to 11. A strategy to obtain transgenic elm based on these results is now in progress.

In vitro micropropagation of Arctostaphylos uva-ursi (L.) Sprengel.: Comparison between two methodologies

In vitro micropropagation of Arctostaphylos uva-ursi was performed to increase the number of ground cover species able to serve as substitute for members of the Rosaceae susceptible to fire blight. Explants (node segments) excised from plants growing in the greenhouse were established in vitro on a medium containing 10 μM α-naphthaleneacetic acid (NAA) and activated charcoal (2 g I-1). Using in vitro grown shoots, two propagation procedures were used:- Culture of nodal fragments with 50 μM NAA resulted in the growth of 6 to 7 nodes every 4 weeks, yielding 1 700 almost rootable shoots after 4 subcultures;- Development of axillary shoots obtained with media containing 25 μM benzyladenine (BA) and 20 μM indoleacetic acid (IAA) yielded almost 500 rootable shoots after 4 subcultures. The rate of propagation decreased after the 3rd subculture.Percentage of in vitro rooted shoots reached 98% with diluted micronutrients and 10 μM NAA but 31% of the plants died during acclimatization.

Effects of temperature and hypoxic atmosphere on preservation and further development of in vitro shoots of peach (‘Armking’) and peach × almond hybrid (‘GF 677’)

Abstract To improve planning of in vitro plant production and/or to preserve germplasm, micropropagated shoots of peach (‘Armking’) and peach × almond (‘GF 677’) were submitted to low oxygen concentrations (0.25-0.2%, plus air as control) combined with decreasing temperatures (25°C, 12°C, 0°C). This method is frequently used in post-harvest physiology to slow down the senescence processes of organs. ‘Armking’ shoots survived for at least 8 months and ‘GF 677’ for 10 months at 0°C. At this temperature, hypoxic conditions had no effect on survival but enhanced regrowth of the shoots afterwards, leading to the maintenance of the multiplication rate. At 25°C and 12°C, hypoxic regimes replaced low-temperature storage, but for shorter times: 2 months for ‘Armking’ and 4 months for ‘GF 677’ at 25°C and at least 4–6 months for the two genotypes at 12°C. This method should thus be used for in vitro preservation of cold-susceptible plants. In addition, hypoxia increased shoot branching capacity, and could be used to promote in vitro micropropagation of species with strong apical dominance.

Multiplication végétative in vitro de Gardenia jasminoïdes Ellis: In vitro Vegetative Multiplication of Gardenia jasminoïdes Ellis.

The vegetative propagation in vitro of Gardenia jasminoides Ellis, an ornamental shrub producing secondary metabolites useful in perfumery and pharmacy, has been investigated. In the presence of benzylaminopurine (0.3 to 1 mg l(-1)) and indoleacetic acid (1 mg l(-1)), axillary shoots arose from nodes of greenhause plants, but a decrease of morphogenic activity was noticed in subcultures. Mother plants have been developed in vitro from which shoots may be taken with the most effective system being a monthly subculture of nodal explants. As many as one million shoots per year could, theoretically, be produced from one shoot grown in vitro. Rooting occurred easily in three weeks in vitro in the presence of indoleacetic acid (1 mg l(-1)) and charcoal (2 g l(-1)) or in vivo by soaking the base of the shoot in the same auxin for two hours. After acclimatization, the rooted plants developed and flowered normally. The successive stages have been sufficiently well monitored to be used as an effective means of vegetative propagation.

Physiological state and clonal variability effects on low temperature storage of in vitro shoot cultures of elms (Ulmus sp.)

As a means of protecting elms against Dutch Elm Disease (DED), in vitro storage of shoot cultures of some elm genotypes under reduced temperatures was investigated. At 7°C under 8 h of fluorescent lighting, survival rate remained high (> 82%) during the storage periods (24–30 months), provided the shoots used for the storage were already rooted or about to root (1–2 months). Necrosis of some axillary ramifications at the end of the storage period reduced the multiplication rate from 3–4 to 1. According to the clone, explants taken from the stored plantlets were not as easy to root as the control (20–30% for U. campestris, OCBi). In order to store elm clones free of DED and to reduce in vitro transfers, an optimal storage period of 18–24 months could be retained. It may be possible to extend the period by improving rooting and growth during the pre-culture phase.