Liquid Culture Systems for in vitro Plant Propagation

418 J.P. Ducos et al.
their weight by an upward airflow. The airflow rate was progressively
increased unit by unit (1 minute for each unit). At each step, the embryo
population was collected at the exit of the column. The number of embryo
per class size was measured using an image analysis system (software
Hellix, Microvision Instruments)
2.4 Sowing
Dehydrated embryos were placed in the compartimented hopper of a
precision seeding system operating by suction (model Step-O-Mat, Visser).
This apparatus uses nozzles to pick up the seeds from the vibrating hopper
and deposits them down tubes into individual cells of the horticultural trays.
The diameter of the nozzles was 0.25 mm. The seeder was adjusted to
distribute the embryos onto polystyrene trays, each one containing 240 cells
filled with soil. The experiments were carried out in a non-sterile
environment (temperature: 24°C; R.H.: 25-30 %).
2.5 Germination
Carrot embryo germination was achieved according to Dupuis et al.
(1999): i) somatic embryos were grown on cellulose acetate miniplugs
moistened with a sterile liquid germination medium ii) a phytoprotection
mixture against micro-organisms was added to this medium as the
experiment was carried out in non-sterile conditions. This mixture contained
a fungicide (iprodione 100 mg l -1 ) and an antibiotic (cefotaxime 100 mg l -1 )
(Dupuis et al., 1999). Each miniplug was placed into an individual glass jar.
The germination medium was composed of macronutrients of Heller (1953),
micronutrients of Murashige and Skoog (1962) and 15 g l -1 sucrose. The
conversion rate was defined as the percentage of embryos giving rise to
normal plantlets after 21 days. The plantlets were characterized by the
initiation of at least two leaves and growth of the root.
3. Results and discussion
3.1 Sorting
Bulk populations of dehydrated somatic embryos were collected by
gently tapping the paper disks held vertically then placed into a seed selector
consisting of an air column. The higher the airflow rate, the longer the
embryos collected at the exit of the air column. Starting from a bulk