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3 AUTHORS:
Rosna Mat Taha
University of Malaya
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Asmah Awal
Universiti Teknologi MARA
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synthetic seeds were encapsulated with 3% sodium alginate with the addition of MS basal
(free calcium), 3% sucrose and 2.0 mg/L BAP, and 0.5 mg/L NAA (explant germinating
medium). The germination rate of the synthetic seeds significantly reduced for both micro
shoots (56.01.1) and somatic embryos (41.41.0) when the encapsulation matrix only
contained 3% sodium alginate with the addition of MS basal (free calcium) and distilled
water. The survival rates also reduced significantly. Synthetic seeds of Begonia x
Hiemalis Fotch. produced 90.48% germination rate (Asmah et al., 2007) when 3%
sodium alginate was used as the encapsulation matrix. The highest germination frequency
of encapsulated micro shoots and somatic embryos were then transferred to vials
containing sowing medium. Most of the beads managed to survive after eight weeks in
culture on MS basal medium, 100% (encapsulated micro shoots) and 90% (encapsulated
somatic embryo), sterile garden soil, 90% (encapsulated micro shoots), 83%
(encapsulated somatic embryo) (Table 3). The survival rate of seedlings reached 75%
after they were transferred to the green house. The success of retrieving complete
plantlets in vitro from encapsulated axillary buds or shoot tips on various planting
mediums has been achieved in few herbaceous plants (Sharma et al., 1994) and woody
species (Bapat et al., 1994).
Storage of Synthetic Seeds
The result showed that 75-95% germination rate could be obtained from one to
two months storage at 4C, whereas three months storage also showed precocious
germination and the seeds germinated at 77% (encapsulated micro shoots) and 60%
(encapsulated somatic embryos). The four- to six-month storage seeds showed lower
germination rate (8-50%). In contrast, the germination frequency of encapsulated micro
shoots and somatic embryos after 120 days storage was lower than after 30 days storage
(Table 4). It is thought that the decline in the germination frequency observed among
encapsulated micro shoots and somatic embryos of Gerbera jamesonii sotred at low
temperature may be due to inhibited respiration of plant tissue by alginate (Redenbaugh et
al., 1987). Storage of somatic embryos or shoot buds using alginate encapsulation
protocol has been attempted in a few species with various degree of success (Bapat and
Rao, 1988; Ghosh and Sen, 1994). Fewer than 5% of Santalum album L. (Bapat and Rao,
1988) encapsulated embryos germinated, followed by storage at 4oC for 45 days and only
8.3% of encapsulated Asparagus cooperi baker. somatic embryos germinated. Most of the
artificial seeds developed into normal seedlings. No phenotypic changes were observed
from the plantlets and over 80% of the plantlets developed into healthy plants. Plantlets
germinated from the germination of synthetic seeds of Gerbera jamesonii (Fig. 1e) were
morphologically identical to the mother plant and developed normally and produced
flowers after six months being transplanted to the green house (Fig. 1f). Results showed
that synthetic seed produced via tissue culture system could serve as a mean for a new
plant line produced through biotechnological improvements to be transplanted to the
field. Synthetic seeds produced could also be handled as true seeds. Furthermore, it has
the potential for long term storage without losing its viability.
CONCLUSION
Synthetic seed technology could be a valuable aid to economical large-scale clonal
propagation and germplasm conservation of Gerbera jamesonii Bolus ex. Hook f. The
possibilities of using both embryogenic and non-embryogenic, in vitro derived vegetative
propagules for encapsulation have been successfully reported in this study. Both explants
were successfully encapsulated with 3% sodium alginate with the addition of MS basal
(free calcium), 3% sucrose and 2.0 mg/L BAP, and 0.5 mg/L NAA. The highest
germination rate was 74.52.6 of encapsulated micro shoots and 54.21.3 of somatic
embryos.
94
ACKNOWLEDGEMENTS
This work was partially funded by Vote F grant number PS077/2007B. The
authors would like to thank the University of Malaya, Malaysia for the above financial
support. The authors would also like to thank the Ministry of Science Technology and
Innovation, Malaysia for the Science Fund Grant SF 12-02-03-2037.
Literature Cited
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Begonia x hiemalis Fotch. CATRINA- International Journal of Environmental
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Bapat, V.A., Mhatre, M. and Rao P.S. 1987. Propagation of Morus indica L. (mulberry)
by encapsulated shoot buds. Plant Cell Rep. 6:393-395.
Castillo, B., Smith, M.A.L. and Yadav, U.L. 1998. Plant regeneration from encapsulated
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Fowke, L.C., Atree, S.M. and Pomeroy, M.K. 1994. Production of vigorous desiccation
tolerant white spruce (Picea glauca [Moench] Voss.) synthetic seeds in a bioreactor.
Plant Cell Rep. 13:601-606.
Gosh, B. and Sen, S. 1994. Plant regeneration from alginate encapsulated somatic
embryos of Asparagus cooperi baker. Plant Cell Rep. 13:381-385.
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production from Paulownia elongate. Plant Cell Rep. 22:16-24.
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Pattnaik, S. and Chand, P.K. 2000. Morphogenic response of the alginate-encapsulated
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95
Tables
3.0
+++
+++
+++
++++
++++
4.0
+++
+++
+++
++++
++++
5.0
+++
+++
***
***
***
6.0
+++
+++
***
***
***
Table 2. Growth response of micro shoots and somatic embryo of Gerbera encapsulated
in different encapsulation matrix. Unencapsulated embryos were used as control.
Encapsulation
matrix
Control
Ca-free MS +
distilled water
Ca-free MS +
3% sucrose
Ca-free MS +
3% sucrose +
2.0 mg/L BAP +
0.5 mg/L NAA
56.01.1a
41.41.0a
64.80.7a
46.10.5a
72.10.8a,b
50.61.5a,b
68.91.2a,b
54.51.1a,b
74.52.6c
54.21.3 c
75.00.5c
63.21.0c
Mean SE, n=30. Mean with different letters differ significantly at p=0.05
96
Ca-free MS +
3% sucrose
Garden soil
Vermiculite
25
19
83
63
97
Figures
4.0 mm
5.0 mm
10 mm
10 mm
2.0 mm
5.0 cm
98