Liquid Culture Systems for in vitro Plant Propagation

264 Katerina Grigoriadou et al.
culture is hyperhydricity, a physiological abnormality of cultivated tissues
causing anatomical deformities and failure of shoot development or rooting
(Ziv and Ariel, 1992; Miguens et al., 1993). Different procedures and
devices have been developed including temporary immersion systems (TIS)
designed especially to overcome these problems (Teisson et al., 1996;
Escalona et al., 1999). Many vessels, mainly consisting of more or less
complex containers, have been used for periodical immersion of the
explants. Some of them are commercially available. This technique has been
successfully used for micropropagation of tropical crops such as banana,
pineapple, sugarcane, coffee and rubber tree, while its efficiency for other
woody species including pear and apple rootstocks has been reported to be
promising, providing the advantages of increased proliferation rate and the
possibility of automation (Teisson et al., 1996; Escalona et al., 1999;
Damiano et al., 2000; Welander et al., 2001). Furthermore, the formation of
shoot clusters in disposable presterilized plastic bioreactors for large-scale
micropropagation of plants has recently become a reality as commercially
available devices, easy to set up and operate in any laboratory, have been
already developed by manufacturers involved in micropropagation industry
(Ziv et al., 1998; Peak et al, 2001).
The in vitro culture of olive tree has not been very successful, mainly due
to the problems of poor microshoot formation, which is cultivar dependent
(Dimassi, 1999; Rugini et al., 1999; Grigoriadou et al., 2002). Moreover,
hyperhydricity problems are usual in olive tissues, which accumulate a
mucus material in the intercellular spaces inhibiting the normal development
of the cultures (Grigoriadou, 2003). The use of a TIS might improve
microshoot development during the proliferation phase and overcome
hyperhydricity problems.
The aim of this work was to study the effect of a new TIS, designed in
VITRO HELLAS S.A. laboratory, on olive microshoot formation, in
comparison with other liquid culture techniques and devices successfully
applied to different woody species.
2. Materials and methods
2.1 Plant material and culture conditions
Stock in vitro cultures of the Greek olive tree (Olea europaea L.) cv.
Chondrolia Chalkidikis were maintained in 500 ml glass jars containing 125
ml of Woody Plant Medium (Lloyd and McCown, 1981), supplemented with
20 µmol zeatin, 10 µmol GA3, 0.3 µmol NAA, 2% (w/v) sucrose and 0.5%
(w/v) agar (B&V S.r.L., type S 1000). The pH was adjusted to 5.2 before