Liquid Culture Systems for in vitro Plant Propagation

272 Katerina Grigoriadou et al.
4. Discussion
Olive trees are mainly grown in the Mediterranean basin, where the
average temperature is high during the vegetative period. The development
of new shoots starts usually in May and continues all over summer when the
temperature is occasionally extremely high. Faster development of
microshoots at high temperature (28 o C) compared to 22° C may be
explained due to the adaptation of the olive plants to such an environment.
Nevertheless, quicker degeneration of plant material sometimes could be a
disadvantage for in vitro culture at high temperature.
The use of a liquid medium for in vitro culture is considered quite
advantageous for mass production due to increased proliferation rates
combined with low production cost based on easy handling and the
simplicity of medium replacement (Teisson et al., 1996; Escalona et al.,
1999). Different methods have been developed to increase the efficiency of
liquid culture technique, such as the use of a raft to support plants over
liquid, addition of liquid medium to cultures already established in agar and
mist cultures. Among them, the TIS has been reported to be successfully
used for many species, the majority of which are from tropical regions
(Teisson et al., 1996; Escalona et al., 1999). Some of the TIS are now
available on the market, such as the RITA system (Teisson et al., 1996),
while others have been used only for research in laboratories (Damiano et
al., 2000; Welander et al., 2001).
The novel TIS used in the present study is based on the same principle as
that used in other similar systems but it was designed in such a way that the
compressed air, which moves the piston of the pneumatic cylinder and
controls immersion movement, is prevented from entering the vessel. Thus,
theoretically the possibility of contamination was reduced. In practice,
contamination problems were initially noticed and for that reason micropore
filters were added. The mechanism that controls the movement was adjusted
only in closed vessels but gas exchange could also be easily achieved via
openings at the top of the vessel protected by sterile filters.
This system worked without problems during the period of the
experiments, presenting the advantages of reliability, easy change of the
medium and easy assembling and disassembling for autoclaving, without
these procedures affecting its functioning.
In olive tissue culture, where low propagation rates and slow microshoot
growth seem to be the main problem (Rugini et al., 1999), the effect of
liquid medium using different culture methods was tested aiming at easier
and faster absorption of nutrients by the cultivated tissues. Even though the
percentage of buds developed into microshoots remained high, the main
disadvantage of liquid media culture, namely the hyperhydricity, proved to