1) Synseed technology involves encapsulating plant propagules like somatic embryos, shoot tips, or buds in a gel matrix, usually sodium alginate, to create artificial seeds that can be stored and germinated like real seeds. 2) Various fruit crops have been successfully propagated using synseed technology including mango, banana, apple, grape, guava, and papaya. Somatic embryos and shoot tips are common propagules used. 3) Advantages of synseed technology include maintaining genetic uniformity, facilitating long distance transportation and exchange of plant materials, and providing a means of propagation for hybrids, endangered species, and plants with recalcitrant seeds.

2. SYNSEED TECHNOLOGY IN FRUIT CROPS
SPEAKER
NEEHARIKA KANTH
RESEARCH SCHOLAR
SUPERVISOR
Prof. Anil K. Singh

3. SYNSEED
• Term “artificial seed” coined by
Murashige, 1977.
• Synonyms: Manufactured seed,
synthetic seed or synseed.
• Artificially encapsulated somatic
embryo, shoot, bud or any other
meristematic tissue that can be
used as functional mimic seed for
sowing.
• Possesses the ability to convert into
a plant under in-vitro or ex-vitro
conditions, and can be stored (Ara
et al.,2000).

4. Propagules for synseed
• Somatic embryo
• Non-embryogenic vegetative propagules
apical shoot buds
axillary bud
nodal segments

6. Why SYNSEEDS?
• Heterozygosity of seeds
• Minute seed size
• Presence of reduced
• endosperm
• Low germination rate
• Seedless varieties
• Recalcitrant seeds
• Storage issues
• Extended juvenility
• Germplasm in field gene banks
exposed to natural disasters
• Distribution and exchange from
field gene banks are difficult

8. • Shoot buds cut to 2-3mm in size and placed in
the encapsulation matrix ( sodium alginate )
• Those cultured plant parts allowed to form
somatic embryos

9. • Using a sterile pipette, the shoot bud or
somatic embryo drawn up with encapsulation
matrix

10. • The shoot bud or somatic embryo is dropped
into the solution ( calcium nitrate ) and a
capsule is formed and allowed to harden
• A thin layer of coating agent is used to coat
the encapsulation matrix

13. COATING AGENTS
• Sodium alginate
• Potassium alginate
• Sodium pectate
• Carboxymethyl cellulose

14. Types of synseeds

15. Dessicated
 Decreasing relative humidity
Slowly, period of 1- 2
weeks (for immature
embryo)
Rapidly, by unsealing the petri
dishes and leaving them overnight
to dry (for many mature embryos)

16.  High osmotic potential
 Low temperature
 Nutrient deprivation
• Increased permeating osmoticants,
sucrose, mannitol
• Non-permeating osmoticants, PEG
• High gel strength media

17. • Calcium alginate capsule pretreated
with potassium nitrate becomes soften
and allow the easily emergence of
shoot and root from alginate beads.
• Application of potassium nitrate also
helps in the breaking of alginate.
• After desiccation, embryos coated with a
protective and nutritive layer that inhibits
mechanical damage during handling and
provides nourishment during the early stages
of conversion. Pond and
Cameron, 2003

18. Non-toxic
Non-aqueous
Melt at a low temperature
Able to adhere to embryo
Soft coating.
CHARACTERISTICS OF COATING MATERIAL

19. Hydrated
• Hydrogels
Agar
 sodium alginate
Potassium alginate
Sodium pectate
Gelatin
Gelrite
Guargum

20. Sodium
alginate best
• no toxicity for
propagules
• quick gellation
• low cost.
Sodium alginate
and calcium salt
• Non-damaging
• Low price
• Easy to use
• High embryo-to-
plant conversion.
Rai et al.
(2009)
Saiprasad, 2001;
Swamy et al., 2009

21. Hydration process
Encapsulated propagules cultured on nutrient medium or different substrates
Washing with sterile double distilled water
Replacement of sodium ions by calcium ions forming calcium alginate
Hardening of alginate beads
Incubation in calcium chloride solution (30- 40 min.)
Dropped into a complexing agent such as calcium chloride or calcium nitrate solution
Prepared in double distilled water or liquid nutrient medium
Mixed with encapsulation mixture [sodium alginate}
Propagules carefully isolated from in vitro cultures

22. Hydrogel encapsulation techniques
Single
layered
Double
layered
Hollow
beads

23. Single layered
• Simplest way to achieve hydrogel
encapsulation.
• Propagules are carefully isolated either from
an in vitro or an in vivo source and mixed with
a suitable hydrogel such as sodium alginate.
• Solution dropped along with the propagule, as
a bead into a agent, calcium chloride or
calcium nitrate .

24. • The pH of both the alginate matrix and complexing
agent adjusted to 5.8.
• The major principle is the formation of round and firm
calcium alginate beads due to an ion exchange process
between sodium and calcium ions.
• The solution in which beads are formed is constantly
stirred/shaken in order to avoid them sticking to each
other and to increase the formation of spherically-
shaped beads.
• Beads placed on a nutrient medium or sown in
different planting substrates.

25. Double layered synseed
• Single layered bead is further coated with a
sodium alginate solution and dropped into
calcium chloride solution for 30 min, then
washed with DDW.
• These double-encapsulated synseeds have all
the merits of a single-layered synseed,with
the added advantage of double encapsulation
for better protection.
Pinker and Abdel-
Rahman (2005)

26. Hollow beads
• Propagules suspended into a solution
containing carboxymethyl cellulose (CMC) and
calcium chloride.
• Dipped into a constantly-stirred sodium
alginate solution.

27. PROPAGULES FOR SYNSEED
PRODUCTION
Bipolar propagules
(Somatic embryos)
Unipolar
propagules

29. Plant species Propagules encapsulated
Ananas comosus Axillary buds
Carica papaya Somatic embryos
Malus pumila (Apple rootstock M 26) Apical and axillary buds, Nodes, Shoot tips
Mangifera indica Somatic embryos
Musa sp. Shoot tips, Shoot apices, Somatic embryos
Psidium guajava Somatic embryos, Shoot tips, Nodal
segments
Punica granatum Nodal segments
Vitis vinifera Somatic embryos

30. Apple
• Different explants of Apple
rootstock M.26 (Malus pumila)
were encapsulated.
• As compared to non-encapsulated
M.26 apple rootstock, single nodes
encapsulated in alginate beads
immediately after a 24-hour
treatment with IBA and sucrose in
dark produced much lower
percentage of plantlet conversion.
• Addition of growth regulators to
the artificial endosperm and culture
of the single nodes for root
primordia initiation allowed
production of comparatively higher
percentage of plantlet conversion.
Sicurani et al. (2001), Brischia et al., 2002

31. Banana
• Shoot tips isolated from multiple shoot
cultures of banana cv. Basrai were
encapsulated in 3% sodium alginate
containing different gel matrices.
• The encapsulated shoot tips regenerated
in vitro on different media as well as
substrates such as filter paper and cotton.
• Cultivars with the B genomes exhibited
better response.
• Rasthali (AAB genomic group) good to
produce synthetic seeds.
• Maximum conversion into plantlets from
encapsulated embryos was achieved
when somatic embryos encapsulated in
5% sodium alginate and cultured on MS
basal medium.
Suprasanna et al.,
2001, Ganapati et al.,
2001

32. Grape
• Cotyledonary-stage somatic embryos originating
from leaf explants were encapsulated individually
in 2% alginate gel.
• encapsulated somatic embryos converted into
plantlets successfully on 0.7% agar medium
containing MS microsalts and 3% sucrose.
• Transferring the embryos onto MS medium prior
to encapsulation resulted in extended storage of
up to 90 days without loss of the conversion
potential.
Das et al., 2006

33. • Encapsulation of individual
somatic embryos and
encapsulated shoot tips.
• Most of encapsulated somatic
embryos (about 90%) remained in
quiescent state similar to
dormancy of zygotic embryos.
• Encapsulated somatic embryos
converted into plantlets after
transfer to medium containing 3%
sucrose.
• The temporary suppression in
conversion of encapsulated
somatic embryos by ABA or high
sucrose offers a possibility of
conservation of elite genotype of
guava for short-period.
GUAVA
Rai et al., 2008

34. Plant regeneration from encapsulated shoot tips of guava. (A) Shoot tips encapsulated in
Ca-alginate beads. (B) Shoot and root emergence from alginate-encapsulated shoot tip.
(C) Plantlet regeneration on agar-solidified MS medium. (D) Plantlet regeneration on MS
medium. (E)Well-developed plantlets regenerated fromencapsulated shoot tips.

35. Mango
• Somatic embryos were
encapsulated in 2% sodium
alginate solution.
• The encapsulated somatic embryos
converted into plantlets
successfully on MS medium.
• The percentage of conversion of
encapsulated somatic embryos was
higher than that of naked somatic
embryos of the same size on the
same medium.
Krishna and Singh, 2007

36. Papaya
• Suitable candidate for
encapsulation technology.
• Cotyledonary stage somatic
embryos encapsulated in
sodium alginate and calcium
chloride to produce synthetic
seeds in papaya.
• 2.5% sodium alginate
prepared in liquid half-strength
MS medium and relatively
short exposure to calcium
chloride were optimum.
Castillo et al. (1998)

37. Pineapple
• Axillary buds isolated were encapsulated in 3%
sodium alginate.
• Maximum conversion of synthetic seeds to
plantlets occurred.
• Synthetic seeds stored at 4 °C remained viable
without sprouting for up to 45 days.
• Encapsulation of microshoots
• in alginate beads for short-term storage of
pineapple.
Gangopadhyay et al. (2005)

38. A. Micro shoots encapsulated in sodium alginate beads.
B. ‘Conversion’ – Rupturing of beads showing proliferation of
micro shoots
after storage.
C. Fully grown-hardened micropropagated plant in liquid
medium.

39. Pomegranate
• Encapsulation of in – vitro shoot
culture using calcium alginate
hydrogel containing MS medium
supplemented
• A combination of 3% sodium
alginate and 100 mM
• calcium chloride was most
suitable for formation of ideal
synthetic seeds.
• Of the different planting media
evaluated, percent sprouting
(shoot development) was
highest in Encapsulated nodal
segments.
Naik and Chand
(2006)

40. Advantages
 Hybrid plants can be easily propagated using synthetic
seed technology.
 Genetically modified plant or crops can be propagated
using synthetic seed technology.
 Endangered species can be propagated using synthetic seed
technology.
 Elite genotype can be preserved and propagated using
artificial seed technology.
 Synthetic seed production is cost effective when compared
to traditional method.

41.  Synthetic seeds can be directly used in fields.
 Genetic uniformity is maintained by using
synthetic seed technology.
Synthetic seeds can be transported from one
country to another without obligations from
quarantine department.
 Synthetic seed transportation is easy as these do
not contain any disease causing agents.
Synthetic seeds are small therefore they are easy
to handle.

42.  Synthetic seed encapsulation provides aseptic
condition to the plant material or explants ,
which is present inside the capsule.
 Herbicides can be added to the formulation .
Synthetic seed crops are easy to maintain
because of uniform genetic constituent.
Synthetic seeds can be made available
throughout the year whereas most of tree plants
produce seeds only in certain months of the year.

43. Limitations
• Hindered emergence of the root or shoot of
the encapsulated propagule by the gel
capsule.
• Hydrated synseeds are sticky and difficult to
handle on a large scale and dry rapidly in the
open air.
• Low soil survival.
• Needs technical skill.

44. Future prospects
• Offers tremendous scope for the conservation
and germplasm exchange of several fruit plants.
• Standardization of methods for synchronization
of developing propagules followed by automation
of the whole process of sorting, harvesting,
encapsulation and germination of the coated
propagules can enhance the pace in the
production of synthetic seeds.