Liquid Culture Systems for in vitro Plant Propagation

368 Barbara Ruffoni et al.
3. Results
After 30 days of culture 60 % of the excised meristems showed asepsis.
Vegetative development was achieved from 23 % of these aseptic explants.
Continuous shoot proliferation was achieved after 2 months in culture in
semisolid medium; in some cases white-reddish callus was recorded at the
base of the shoots.
From the explants transferred in liquid medium, biomass proliferation
was obtained with both genotypes tested and some differences were
recorded. In the cv. PAN, after the first subculture (30 days) the biomass
weight values were similar in all treatments. At the end of the second
subculture (60 days), in the presence of either 2.22 and 4.44 µmol BA, the
fresh weight was at least doubled and significantly increased with respect to
the control treatment. The addition of ANC seemed to inhibit biomass
proliferation (Figure 1). After transfer to the hormone-free medium, the
biomass production of the cv. PAN remained low for the cultures coming
from ANC-supplemented medium, while the fresh weight of the cultures
coming from 4.44 µmol BA increased significantly (Figure 2).
In the cv. RE, the callus showed an homogeneous behaviour among the
treatments both during culture in the presence of growth regulators and after
the transfer to hormone-free medium. With respect to the control treatment,
greater fresh weight biomass increase was recorded for the cultures coming
both from 2.22 and 4.44 µmol BA; the previous presence of ANC did not
affect this proliferation (Figure 2).
In the cultures supplemented with BA or BA+ANC, the primary shoot
explants grew, but also differentiated yellowish or brown/red hard callus at
their bases. In the genotype RE, in the medium containing 2.22 µmol
BA+ANC, a large number of bulblets grew per unit of biomass (callus)
(Figure 3). This difference was not statistically confirmed.
In the cultures of both genotypes transferred to media lacking growth
regulators, several shoot primordia and complete small plantlets developed
from the callus and the cell aggregates.
The mean comparison of the data related to the shoot development in
growth regulator-free medium is reported in figure 4. The response of the
two genotypes was similar: the previous presence of ANC strongly inhibited
shoot development and the greatest number of shoots was observed in
cultures previously grown with 2.22 BA (7.25 shoots per callus unit).