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Rice Genomics and Genetics 2013, Vol. 4, No. 4, 14-27
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18
3.2.4 Plant growth regulators
To improve the storability and germination of
synthetic seed, many workers used growth regulators.
Encapsulation with GA3 was found to be useful for
storage of somatic embryos of citrus at 4
℃
for 1
month (Mariani, 1992; Antonietta et al., 1998). Buds
treated with 0.01~1.0 mM ABA either prior to
encapsulation or even in the alginate, matrix to inhibit
the precocious growth (Palmer and Jasrai, 1996).
Synthetic seeds obtained from the excised embryos of
intact seeds treated with higher (2~3 mg/L)
concentrations of ABA showed tolerance to low
temperature storage and retained higher germination
percentage. ABA is implicated as a controlling factor
for germination and dormancy in somatic embryos
and seeds (Senaratna et al., 1995). Ruffoni et al. (1994)
found that the addition of zeatin (0.5 mg/L) to the
alginate encapsulation coating improved the shoot
production and sucrose (40 mg/L) added to MS agar
medium improves the percentage root emergence of
Enstoma grandiflorum.
Desiccation tolerance is a characteristic of somatic
embryos that must be induced and therefore, require a
pretreatment with ABA or stress elicit the desired
response. The importance of ABA application for
imparting desiccation tolerance during storage of
somatic embryos was well recognized (Senaratna et al.,
1989; Senaratna et al.,1990; Lecouteux, 1993). Kitto
and Janick (1985) reported that ABA effectively
hardened carrot embryos, permitting them to survive
desiccation. In a further study, Kim and Janic (1989)
found that ABA at 10-6 M effectively increased
embryo survival after desiccation suggesting that ABA
hardens somatic embryos of celery. They also
achieved desiccation tolerance by a combination of
ABA and proline. ABA increased the accumulation
and altered the distribution of fatty acids in somatic
embryo. Takahata et al., (1992) reported that the
desiccated embryos lost their viability if not treated
with ABA. Dormancy and desiccation tolerance can
be imparted to the embryos by treating them with
appropriate concentration of ABA or high sucrose
concentrations (Anandarajah and McKerrie, 1990).
The ability of somatic embryos to with stand to low
moisture content is important for storage, and also
plays a big role in developmental transition between
maturation and germination. Phokajornyod et al.
(2004) found that somatic embryos that treated with
0.5 mg/L of ABA for 20 days before encapsulation
with sodium alginate, and dehydrated in laminar flow
hood until 80% water loss still remained germinated
to 58%. Furthermore, improvement of dry somatic
embryos was accomplished by adding 60 g/L sucrose
in the maturation medium which resulted 40% of
plantlets conversion after four weeks of storage in
ambient temperature.
The germination percentage of artificial seeds of
Brassica oleracea var. botrytis was enhanced by the
inclusion of 0.3 mg/L NAA and 3.0 mg/L BAP in the
encapsulation matrix after 7 and 30 days of
pregermination storage (Siong et al., 2012). The time
taken for germination was also faster (5 days after 7
days of storage and 11 days after 30 days of storage)
when MS fortified with 0.3 mg/L NAA and 3.0 mg/L
BAP were used. Results Zhang et al. (2011) showed
that maltose, active carbon and sodium alginate played
the important role in germinate of D. Candidum
artificial seeds. The optimal condition was maltose 4%,
hormone rate between 6-BA and NAA 12:1, active
carbon 0.3%, sodium alginate 4%, time of ion
exchange is 5 min for artificial seeds procedure.
3.2.5 Use of protective chemicals and microorganisms
Protective chemicals may be used to protect the
synthetic seed from fungal and/or bacterial infection.
To avoid bacterial contamination, Ganapathi et al.
(1992) added an antibiotic mixture (0.25 mg/L)
containing rifampicin (60 mg), cefatoxime (250 mg)
and tetracycline-HCl (25 mg) dissolved in 5 ml
dimethyl sulphate to get matrix. Mycorhizas,
pesticides and fertilizers can be incorporated into the
capsules to enhance germination rates and seedling
growth (Toruan and Sumaryono, 1994). Sharma et al.
(2000) used mycorrhiza for preparation of synthetic
seeds. Activated charcoal (0.1%) can also be added to
the matrix to absorb the polyphenol exudates of the
encapsulated shoots of banana (Ganapathi et al.,
1992).
Zhang et al. (2001) prepared synthetic seeds of
Dendrobium candidum using clay and vermiculate