Protocols for Micropropagation of Woody Trees and Fruits

MICROPROPAGATION OF VACCINIUM VITIS-IDAEA L. 461
recorded in V. vitis-idaea. On media supplemented with 2.19 mg l –1 zeatin callus
formation started on leaf explants of cultivar ‘Red Pearl’. After transfer of callus on
AN medium with 0.5 mg l –1 zeatin (Table 1) adventitious buds began to appear on
the callus surfaces. After 3 subcultures the number of adventitious shoots regenerated
from the leaf-originated callus was considerable high and reached number 122.46
shoots per explant (Figure 1B,C).
2.3.3. Influence of Medium pH on Micropropagation of Vaccinium vitis-idaea L.
It is known that by changing pH values nutrient availability and plant metabolism
are affected. Vaccinium spp. production is limited to soils with naturally low pH or
soils treated with acidifying amendments (Finn et al., 1991). For understanding of
pH tolerance in vitro screening with modification of pH values in AN medium with
0.5 mg l –1 zeatin within a tested pH range 4.0–6.0 was performed in selected cultivars
of V. vitis-idaea cvs. ‘Red Pearl’ and ‘Koralle’ during shoot formation from apical
and axillary buds (Ostrolucká et al., 2004).
In cv. ‘Koralle’ with increasing pH values in the range 4.0–5.5 the increasing
tendency in shoot proliferation intensity was observed during 1–3 subcultures, with
the highest shoot number at the third subculture on the medium with pH 5.5 (9.63
shoots/explant). At higher medium pH 6.0, the considerable decreasing in shoot
number was noted (4.63 shoots/explant).
The higher tolerance to medium pH exhibits cv. ‘Red Pearl’, where the highest
value in shoot number was noted at pH 4 (7.83 shoots/explant). In the pH range 4.5–
5.5 the number of formed shoots was similar (4.73–5.10), while at medium pH 6 the
number of shoots declined to 3.13. We can state that cv. ‘Red Pearl’ is much more
tolerant to the substrate pH, what is favourable for the growing in the less demanding
natural conditions. In general, the medium pH value in the range 4–5.5 may be
recommended for micropropagation of lingonberry.
2.3.4. Shoot Proliferation
For long-term proliferation of in vitro regenerated axillary or adventitious shoots
AN medium supplemented with 0.5 mg l –1 zeatin was successfully used. On this
medium formation of vigorous multiple shoot cultures was observed with different
intensity of shoot proliferation in tested cultivars. Increasing of shoot proliferation
intensity can be achieved by segmentation of regenerated, elongated microshoots on
one-node segments and their further cultivation on medium with 0.5 mg l –1 zeatin
(Figure 1D).
2.4. Rooting and Hardening
Satisfactory rooting of isolated microshoots, alike in Vaccinium corymbosum, is
possible to achieve by two approaches:
1. ex vitro rooting – by short (2–3 min) dipping of 15–20 mm long microshoots
into 0.8 mg l –1 IBA solution followed by direct planting in containers 50
mm in diameter filled with peat substrate;