Liquid Culture Systems for in vitro Plant Propagation

300 M. Vágner et al.
mature somatic embryos per g FW tissue
110
100
90
80
70
60
50
40
30
20
10
0
C111 C109 C110 AFO M22 W2 C105 C107 W1 SC13 L2 C203
cell line
protocol 1
protocol 2
Figure 3: The yield of mature Norway spruce somatic embryos grown on membrane rafts.
Protocol 1: Cultures transferred from proliferation medium (containing 5 �mol 2,4-D, 2 �mol
kinetin and 2 �mol BAP) directly to maturation medium (20 �mol ABA).
Protocol 2: Cultures were transferred from proliferation medium to pre-maturation medium
(no PGR, 1 week), and then to maturation medium. Bars indicate SE.
3.3 Use of PEG in maturation medium
PEG is often added to the media for conifer somatic embryo maturation
as a nonpermeating osmoticum, which increases embryo yield and enhances
embryo development and maturation. On the other hand, the use of PEG in
the maturation medium is sometimes criticized. Find (1997) observed PEGrelated
intercellular spaces below apical meristems of somatic embryos.
Bozhkov and von Arnold (1998) reported a PEG-related decrease of
germination frequency and inhibition of post-germinative root growth. In
our experiments, PEG 4000 at low concentration (1.87 % (w/v) increased
embryo yield and shortened maturation phase: higher PEG concentrations
(up to 5 % (w/v)) had even slightly more beneficial effect in these aspects
(Figure 4). Germination frequency of somatic embryos grown on 3.75 %
(w/v) PEG remained unchanged, whereas 5 and 7.5 % (w/v) PEG 4000
slightly inhibited germination (Figure 5). A marked difference in sensitivity
to PEG treatment was observed between different cell lines. The study of
microscopic sections revealed no morphological aberrations of developing
somatic embryos, which were grown on medium containing 3.75 % (w/v)
PEG, and the resultant germinating plantlets. However, we did not study
development of plantlets after 8 weeks of germination. Thus we cannot