Symbiotic Fungi: Principles and Practice (Soil Biology)

116 H. Dupré de Boulois et al.
16/8 h photoperiod. The photosynthetic photon flux should comprise between
200 and 400 mmol m 2 s 1 .
6. Medium is regularly added (about 5 ml once every 2 to 4 weeks) to the RC to
guarantee plant and fungal growth. The medium is added at a temperature of
40 C in order to reduce the risk of damaging the roots and AM fungi.
7. When an extensive ERM is reached, liquid or solid medium is added into the
HC to ensure mycelium growth in this compartment (see Sect. 7.4.1).
8. Isotopic labelling and harvest are described in Sects. 7.6.3 and 7.6.5.
Remarks
Depletion of the Medium
Due to the transpiration of the host plant, depletion of the medium in the RC occurs
while the plant is actively growing. However, contrarily to the HAM–P in vitro
culture system, the depletion of the medium is limited. Indeed, as the whole plant is
maintained under in vitro conditions, the transpiration of the plant is highly
reduced. It is nonetheless necessary to regularly check the level of medium in the
AM–P in vitro culture system and, even if no medium depletion is visible, medium
should be added regularly to provide nutrients to the plant and the AM fungus. It is
thus advised to add medium depending on the nutritional needs of the plant and AM
fungus. Concentrated medium (2–10-times) can also be used (see for instance
Dupré de Boulois et al. 2006) to provide nutrients. If other ventilation membranes
or Petri plate sizes are used, it is necessary to assess medium depletion to ensure that
the extraradical hyphae can cross the partition wall between the RC and HC.
Plant Transpiration
As the plants develop fully in vitro with only a membrane allowing gas exchange, it
is possible that the relative humidity within the AM–P in vitro culture system is
very high and limits plant transpiration. If this is advantageous as medium depletion
in the RC is low, it may have significant impact on plant physiology such as
plant photosynthesis, nutrient translocation from root to shoot or plant abscisic
acid content.
7.6 Labelling with Isotopic Tracers
Isotopic labelling is performed by using an isotopic tracer to observe the movement
of certain elements in chemical, biological, or physical processes. The term tracer is
applied commonly to any stable or radioactive isotope used to trace the course of
non-radioactive elements or biological substances (e.g. proteins, sugars, DNA), and
also radioactive elements such as radionuclide contaminants. The observations may