GENERAL ç ARTICLE

Artificial Seeds and their Applications

G V S Saiprasad

Plant propagation using artificial or synthetic seeds devel-

oped from somatic and not zygotic embryos opens up new
vistas in agriculture. Artificial seeds make a promising
technique for propagation of transgenic plants, non-seed
producing plants, polyploids with elite traits and plant lines
with problems in seed propagation. Being clonal in nature
G V S Saiprasad is a the technique cuts short laborious selection procedure of
scientist in Division of
the conventional recombination breeding and can bring the
Biotechnology, Indian
Institute of Horticulture advancements of biotechnology to the doorsteps of the
Science, Bangalore. He is farmer in a cost-effective manner.
currently working on
molecular events during Need for Artificial or Synthetic Seed Production Techno-
ripening in mango and logy
banana and on molecular
markets. Development of micropropagation techniques will ensure abun-
dant supply of the desired plant species. In some crop species
seed propagation has not been successful. This is mainly due to
heterozygosity of seed, minute seed size, presence of reduced
endosperm and the requirement of seed with mycorrhizal fungi
association for germination (eg. orchids), and also in some
seedless varieties of crop plants like grapes, watermelon, etc.
Some of these species can be propagated by vegetative means.
However, in vivo vegetative propagation techniques are time
consuming and expensive. Development of artificial seed pro-
duction technology is currently considered as an effective and
efficient alternate method of propagation in several commer-
cially important agronomic and horticultural crops. It has been
suggested as a powerful tool for mass propagation of elite plant
species with high commercial value. Characteristics of clonal
propagation systems are discussed in Box 1.

Artificial seed technology involves the production of tissue

culture derived somatic embryos encased in a protective coat-
ing. Artificial seeds have also been often referred to as synthetic

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Box 1. Characteristics of Clonal Propagation Systems

Micropropagation
Low volume, small scale propagation method
Maintains genetic uniformity of plants
Acclimatisation of plantlets required prior to field planting
High cost per plantlet
Relatively low multiplication rate

Greenhouse cuttings
Low volume, small scale propagation method
Maintains genetic uniformity of plants
Rooting of plantlets required prior to field planting
High cost per plantlet
Multiplication rate limited by mother plant size

Artificial seeds
High volume, large scale propagation method
Maintains genetic uniformity of plants
Direct delivery of propagules to the field, thus eliminating transplants
Lower cost per plantlet
Rapid multiplication of plants.

seeds. However, the term ‘synthetic seed’ should not be con-

fused with commercial seeds of a synthetic cultivar which is
defined as an advanced generation of an open pollinated popula-
tion composed of a group of selected inbred clones or hybrids.
The concept of artificial or synthetic seed is shown in Figure 1.

Artificial seed

Somatic embryo

Artificial endosperm

Figure 1. Artificial seed con-

cept.

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 GENERAL ç ARTICLE

Box.2. Advantages of Artificial or Synthetic Seeds over Somatic Embryos for Propagation

Ease of handling while in storage

Easy to transport
Has potential for long term storage without losing viability
Maintains the clonal nature of the resulting plants
Serves as a channel for new plant lines produced through biotechnological advances to be delivered
directly to the greenhouse or field
Allows economical mass propagation of elite plant varieties.

These synthetic seeds would also be a channel for new plant

lines produced through biotechnological advances to be deliv-
ered directly to the greenhouse or field. Advantages of artificial/
synthetic seeds over somatic embryos for propagation are listed
in Box 2. This synthetic seed production technology is a high
volume, low-cost production technology. High volume propa-
gation potential of somatic embryos combined with formation
of synthetic seeds for low-cost delivery would open new vistas
for clonal propagation in several commercially important crop
species.

What are Somatic Embryos?

Somatic embryos are bipolar structures with both apical and

basal meristematic regions, which are capable of forming shoot
and root, respectively. A plant derived from a somatic embryo is
sometimes referred to as an ‘embling’.

Somatic Embryos vs Zygotic Embryos and their

Advantages
Since the natural
Somatic embryos are structurally similar to zygotic embryos
seed develops as
found in seeds and possess many of their useful features, includ-
a result of a sexual
ing the ability to grow into complete plants. However, somatic
process in cross-
embryos differ in that they develop from somatic cells, instead of
pollinating species,
zygotes (i.e., fusion product of male and female gametes) and
it is not genetically
thus, potentially can be used to produce duplicates of a single
identical to one
genotype. Since the natural seed develops as a result of a sexual
single parent.
process in cross-pollinating species, it is not genetically identi-

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cal to one single parent. In contrast, somatic embryo develops

from somatic cells (non-sexual) and does not involve sexual
recombination. This characteristic of somatic embryos allows
not only clonal propagation but also specific and directed changes
to be introduced into desirable elite individuals by inserting
isolated gene sequences into somatic cells. This bypasses ge-
netic recombination and selection inherent in conventional
breeding technology. If the production efficiency and conve-
nience comparable to that of a true seed are achieved, somatic
embryos can be potentially used as a clonal propagation system.
The lack of
synchrony of Basic Requirement for Production of Artificial
somatic embryos Seeds
is, arguably, the
single most Recently, production of synthetic seeds by encapsulating so-
important hurdle to matic embryos has been reported in few species. One pre-
be overcome requisite for the application of synthetic seed technology in
before advances micropropagation is the production of high-quality, vigorous
leading to somatic embryos that can produce plants with frequencies com-
widespread parable to natural seeds. Inability to recover such embryos is
commercialization often a major limitation in the development of synthetic seeds.
of synthetic seeds Synthetic seed technology requires the inexpensive production
can occur. of large numbers of high quality somatic embryos with synchro-
Synchronized nous maturation. The overall quality of the somatic embryos is
embryoid critical for achieving high conversion frequencies. Encapsula-
development is tion and coating systems, though important for delivery of
required for the somatic embryos, are not the limiting factors for development of
efficient production synthetic seeds.
of synthetic seeds.
At present, the characteristic lack of developmental synchrony
in embryogenic systems stymies multi-step procedures for guid-
ing somatic embryos through maturation. The lack of syn-
chrony of somatic embryos is, arguably, the single most impor-
tant hurdle to be overcome before advances leading to wide-
spread commercialization of synthetic seeds can occur. Syn-
chronized embryoid development is required for the efficient
production of synthetic seeds.

42 RESONANCE ç May 2001

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Procedure for Production of Artificial Seeds

Establish somatic embryogenesis

↓
Mature somatic embryos
↓
Synchronize and singulate somatic embryos
↓
Mass production of somatic embryos
↓
Standardization of encapsulation
↓
Standardization of artificial endosperm Alginate hydrogel
↓ is frequently
Mass production of synthetic seeds selected as a
↓ matrix for synthetic
Greenhouse and field planting seed because of
its moderate
Types of Gelling Agents used for Encapsulation viscosity and low
spinnability of
Several gels like agar, alginate, polyco 2133 (Bordon Co.), carboxy solution, low
methyl cellulose, carrageenan, gelrite (Kelko. Co.), guargum, toxicity for somatic
sodium pectate, tragacanth gum, etc. were tested for synthetic embryos and quick
seed production, out of which alginate encapsulation was found gellation, low cost
to be more suitable and practicable for synthetic seed produc- and bio-
tion. Alginate hydrogel is frequently selected as a matrix for compatibility
synthetic seed because of its moderate viscosity and low characteristics.
spinnability of solution, low toxicity for somatic embryos and
quick gellation, low cost and bio-compatibility characteristics.
The use of agar as gel matrix was deliberately avoided as it is
considered inferior to alginate with respect to long term storage.
Alginate was chosen because it enhances capsule formation and
also the rigidity of alginate beads provides better protection
(than agar) to the encased somatic embryos against mechanical
injury. Alginate encapsulated somatic embryos of orchids are
shown in Figure 2 and the plantlets derived from artificial or
synthetic seeds of orchid are shown in Figure 3.

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Figure 2 (left). Artificial or Principle and Conditions for Encapsulation with

synthetic seed produced in Alginate Matrix
orchids by alginate encap-
sulation. Alginate is a straight chain, hydrophilic, colloidal polyuronic
Figure 3 (right). Artificial or acid composed primarily of hydro-β-D-mannuronic acid resi-
synthetic seed derived
dues with 1-4 linkages. The major principle involved in the
plantlets in orchid.
alginate encapsulation process is that the sodium alginate drop-
lets containing the somatic embryos when dropped into the
Somatic embryos CaCl2.2H2O solution form round and firm beads due to ion
lack seed coat exchange between the Na+ in sodium alginate with Ca2+ in the
(testa) and CaCl2.2H2O solution. The hardness or rigidity of the capsule
endosperm that mainly depends upon the number of sodium ions exchanged
provide protection with calcium ions. Hence, the concentration of the two gelling
and nutrition for agents i.e., sodium alginate and CaCl2.2H2O, and the complexing
zygotic embryos in time should be optimized for the formation of the capsule with
developing seeds. optimum bead hardness and rigidity. In general, 3% sodium
To augment these alginate upon complexation with 75 mM CaCl2.2H2O for half an
deficiencies, hour gives optimum bead hardness and rigidity for the produc-
addition of tion of viable synthetic seeds.
nutrients and
Artificial Endosperm
growth regulators
to the Somatic embryos lack seed coat (testa) and endosperm that
encapsulation provide protection and nutrition for zygotic embryos in devel-
matrix is desired, oping seeds. To augment these deficiencies, addition of nutri-
which serves as an ents and growth regulators to the encapsulation matrix is de-
artificial sired, which serves as an artificial endosperm. Addition of
endosperm. nutrients and growth regulators to the encapsulation matrix

44 RESONANCE ç May 2001

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results in increase in efficiency of germination and viability of

encapsulated somatic embryos. These synthetic seeds can be
stored for a longer period of time even up to 6 months without
losing viability, especially when stored at 4 oC.

Addition of Adjuvants to the Matrix

In addition to preventing the embryo from desiccation and

mechanical injury, a number of useful materials such as nutri-
ents, fungicides, pesticides, antibiotics and microorganisms
(eg. rhizobia) may be incorporated into the encapsulation ma-
trix. Incorporation of activated charcoal improves the conver-
sion and vigour of the encapsulated somatic embryos. It has
been suggested that charcoal breaks up the alginate and thus
increases respiration of somatic embryos (which otherwise lose
vigour within a short period of storage). In addition, charcoal
retains nutrients within the hydrogel capsule and slowly re-
leases them to the growing embryo.

Utilization of Artificial Seeds

The artificial seeds can be used for specific purposes, notably In addition to
multiplication of non-seed producing plants, ornamental hy- preventing the
brids (currently propagated by cuttings) or the propagation of embryo from
polyploid plants with elite traits. The artificial seed system can desiccation and
also be employed in the propagation of male or female sterile mechanical injury,
plants for hybrid seed production. Cryo-preserved artificial a number of useful
seeds may also be used for germplasm preservation, particularly materials such as
in recalcitrant species (such as mango, cocoa and coconut), as nutrients,
these seeds will not undergo desiccation. Furthermore, fungicides,
transgenic plants, which require separate growth facilities to pesticides,
maintain original genotypes may also be preserved using so- antibiotics and
matic embryos. Somatic embryogenesis is a potential tool in the microorganisms
genetic engineering of plants. Potentially, a single gene can be (eg. rhizobia) may
inserted into a somatic cell. In plants that are regenerated by be incorporated
somatic embryos from a single transgenic cell, the progeny will into the
not be chimeric. Multiplication of elite plants selected in plant encapsulation
breeding programs via somatic embryos avoids the genetic re- matrix.

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 GENERAL ç ARTICLE

Artificial seeds combination, and therefore does not warrant continued selec-
produced in tissue tion inherent in conventional plant breeding, saving consider-
culture are free of able amount of time and other resources. Artificial seeds pro-
pathogens. Thus, duced in tissue culture are free of pathogens. Thus, another
another advantage advantage is the transport of pathogen free propagules across the
is the transport of international borders avoiding bulk transportation of plants,
pathogen free quarantine and spread of diseases.
propagules across
the international Potential Uses of Artificial Seeds
borders avoiding
Delivery systems:
bulk transportation
of plants, • Reduced costs of transplants
quarantine and
• Direct greenhouse and field delivery of:
spread of
- elite, select genotypes
diseases.
- hand-pollinated hybrids
- genetically engineered plants
- sterile and unstable genotypes
• Large-scale mono cultures
• Mixed-genotype plantations
• Carrier for adjuvants such as microorganisms, plant growth
regulators pesticides, fungicides, nutrients and antibiotics
• Protection of meiotically-unstable, elite genotypes
• Can be conceivably handled as seed using conventional plant-
ing equipment.

Analytical tools:

• Comparative aid for zygotic embryogeny

• Production of large numbers of identical embryos
• Determination of role of endosperm in embryo development
and germination
• Study of seed coat formation
• Study of somaclonal variation.

46 RESONANCE ç May 2001

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Applicability and Feasibility of Artificial Seed

Production Technology

In order to be useful, synthetic seed must either reduce produc-

tion costs or increase crop value. The relative benefits gained,
when weighed against development costs, will determine whether
its use is justified for a given crop species. Considering a
combination of factors, including improvement of the existing
embryogenic systems, relative cost of seed as well as specific
application for synthetic seed allows judgement of relative need
for a given crop. For example, synthetic seed of seedless water-
melon would actually cost less than conventional seed, provid-
ing a benefit at the outset of crop production. Although embryo-
genic systems for this crop do not exist, the benefit that could be
conferred by use of synthetic seed would be very great. Value-
added aspects that would increase crop worth are numerous and
include cloning of elite genotypes, such as genetically engi-
neered varieties, that cannot produce true seed.

Suggested Reading

[1] K Redenbaugh, (ed.), Synthetic seeds, CRC Press, Boca Raton, 1993.
[2] J A Fujii, D T Slade, K Redenbaugh, and K A Walker, Artificial seeds Address for Correspondence
for plant propagation, Trends in Biotechnology, 5, 335-339, 1987. G V S Saiprasad
[3] I Kinoshita, The production and use of artificial seed, Research Journal Division of Biotechnology
of Food and Agriculture, 15 (3), 6-11, 1992. IIHR, Hessarghatta Lake Post
[4] T Senaratna, Artificial seeds, Biotechnology Advances, 10, 379-392, Bangalore 560 089, India.
1992.

There are grounds for cautious opti-

mism that we may now be near the end
of the search for the ultimate laws of
nature.

Stephen W Hawking
A brief history of time:
from the Big Bang to Black Holes

RESONANCE ç May 2001 47