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Endosperm culture and somatic embryogenesis
• TOPIC:
• TRIPLOID PRODUCTION (ENDOSPERM
CULTURE)- DIFFERENTIATION AND FACTORS
AFFECTING THEM- APPLICATIONS
• SOMATIC EMBRYOGENESIS- FACTORS
AFFECTING, STAGES OF SOMATIC
EMBRYOGENESIS, LARGE SCALE PRODUCTION,
SYNTHETIC SEEDS CONCEPT AND ITS PRO’S
AND CON’S
PRESENTED BY
ZUBY GOHAR ANSARI
TAM/14-26
ENDOSPERM CULTURE (HISTORY AND DEF.)
• Double fertilization occurs in majority of
angiosperms which results in two fusion
products i.e. Zygote (fusion product of egg and
one of the male gamete) and triploid endosperm
(fusion product of polar nuclei and second male
gamete).
• The endosperm lack many organogenic potential
and vascular differentiation.
• Endosperm is unique tissue, firstly in its function
of supplying nutrition to developing embryo and
secondly in being triploid.
• Attempts to grow endosperm tissue in cultures
began in 1930’s and now mature and immature
endosperm of various taxa of angiosperm have
been grown.
• Lampe and Mills (1933) : They were first to
report the proliferation of immature endosperm
tissue of maize grown on medium containing
extract of potato.
• Nakano et al. (1975): Successfully cultured
immature endosperm of rice and achieved
organogenesis.
• Rangaswami and Rao (1963): Successfully
cultured mature endosperm of santalum
album (chandan) up to callus.
• Endosperm culture: It is the in vitro
development of isolated mature or immature
endosperm from seed at proper stage on a
suitable culture medium to obtain triploid
plantlet.
Endosperm culture and somatic embryogenesis
TYPES OF ENDOSPERM CULTURE
• 1. Mature endosperm culture: It is the in vitro
development of isolated mature endosperm
from ripen endospermic seed on suitable
culture medium to obtain triploid plantlet is
called mature endosperm culture.
• 2. Immature endosperm culture: It is the in
vitro development of isolated immature
endosperm isolated at precise stage from
immature seed, cultured on suitable culture
medium to obtain triploid plantlet is called
immature endosperm culture.
STEPS IN ENDOSPERM CULTURE
• It consist of mainly 3 steps:
• 1. The immature or mature seeds are dissected
under aseptic conditions and endosperms
along with embryos are excised.
• 2. Excised endosperms are cultured on a
suitable medium and embryos are removed
after initial stage.
• 3. Callus followed by embryogenesis or shoot
bud differentiation.
• 4. Complete plant formation.
Endosperm culture and somatic embryogenesis
PROCEDURE OF ENDOSPERM CULTURE
• 1. Explant source: In most of the cereals, mature
endosperm does not respond to cultural conditions. It is
therefore, excised at proper period of development.
Normally the endosperm of cereals undergo certain
changes 12 days after pollination making this able to
respond. Proper stage for maize is 8-11 days after
pollination, 4-7 days after pollination for rice, and 8 days
after pollination for wheat.
• In some families association of embryo tissue in initial
stages seems essential for proliferation of mature
endosperm. In such cases, entire seeds are used as
explant. Immature seeds provide explant for non-
endospermic seeds.
• 2. Inoculation of explant: For in vitro culture of
mature endosperm, seeds having massive endosperm
are decoated, surface sterilized with suitable
disinfectant and after 2-3 washings with sterile distilled
water, planted on the nutrient medium. For in vitro
culture of an immature endosperm, the entire seed or
kernel is surface sterilized and the endosperm tissue is
excised under aseptic conditions. In cereals, top of
kernel or immature ovaries (micropyle end) are cut off
with a sterile knife and exposed endosperm squeezed
out and placed on the callus induction nutrient
medium.
• The basal white and MS medium is generally used to
induce callus from an immature endosperm.
Endosperm culture and somatic embryogenesis
• The basal medium is supplemented with
tomato juice, yeast extract, grape juice.
Sucrose (2-4%) is used as a source of
carbohydrates. In some cases, addition of 2,4-
D or IAA, cytokinin necessary.
• 3. Incubation/ maintenance of culture: To
induce callus, the endosperm cultures are
maintained darkness or diffused light.
Differentiation take place when calli are
transferred to bright light (2000 lux – 4000
lux) and 25⁰C temperature
• 4. Shoot bud differentiation or
embryogenesis: Plantlet formation from
endosperm callus maturation follow organogenic or
embryogenic mode of development. Embryo
differentiation occurs when the proliferated tissue is
transferred from callusing to basal medium with or
without gibberellins. So culturing carried out up to
complete development of plantlet.
• 5. plantlet developed are hardened in green house
by transferring in vermiculture media and
maintaining proper humidity.
FACTORS AFFECTING ENDOSPERM CULTURE
• 1. Explant stage: proper stage may vary from
cellular (immature) to mature endosperm
depending upon species. (4-7 DAP in rice, 8
DAP in wheat and 12 DAP in maize and fully
matured in santalum album).
• 2. Nutrient medium: Low amount of reduced
nitrogen is required for proliferation.
Undefined source like tomato juice, yeast
extract, etc.
• 3. Physical factors: The pH 7.0 seems to be
effective for fresh weight increase. Maximum
growth of endosperm occurs between 24-
27⁰C temperature and 12-16 hours
photoperiod with diffuse day light supported
callusing as well as regeneration.
• 4. Embryo factors: association of embryo
tissue in the initial stages seems essential for
inducing proliferation of mature endosperm
tissue in some families. Immature seeds of
non endospermic seeds exhibit no
dependence on embryo factor.
APPLICATIONS OF ENDOSPERM CULTURE
• Techniques of endosperm culture has enabled
the production of triploid plants. Triploid
plants are self sterile and usually seedless. The
trait increases edibility of fruits and is
desirable in plants such as apple, banana,
grape, watermelon and mango which are
commercially important.
• In timber and fuel yielding plants, triploids
show better performance over their relative
diploids or tetraploids. Also there is no
problem of seed sterility as they can be
multiplied by vegetative means.
SOMATIC EMBRYOGENESIS
• Regeneration of embryos from somatic cells, tissues
or organs either denovo or directly in vitro conditions
is known as somatic embryogenesis.
• It is also known as non- zygotic/ non-sexual
embryogenesis.
• Various terms for non-zygotic embryos have been
reported such as,
• A. Adventive embryos: Somatic embryos arising
directly from other organs or embryos.
• B. Parthenogenic embryos: Formed by
unfertilized eggs.
• C. Androgenetic embryos: Formed by male
gametophyte.
• Occurence of asexual embryogenesis is
generally restricted to intraocular tissues.
Endosperm culture and somatic embryogenesis
• Differences between somatic and
sexual embryos
SOMATIC EMBRYOS SEXUAL EMBRYOS
Embryos arises from single cell. Arises from multi cell.
Embryos have bipolar structure. It is monopolar structure.
Embryos has no vascular connections with
cultured explant.
Embryos has vascular connections with
cultured explant.
Induction of somatic embryogenesis
requires single hormonal signal.
Requires two hormonal signals.
• The initiation and development of embryos
from somatic tissues in plant culture was first
recognized by steward et al (1958) and Reinert
(1958- 59) in cultures of Daucus carota
(carrot).
• In addition to the development of somatic
embryos from sporophytic cells, embryos have
been obtained from generative cells, such as
the classic works done by Guha and
Maheswari (1964), with Datura innoxia
microspores.
TWO ROUTES TO SOMATIC EMBRYOGENESIS
• Sharp et al (1980) divided 2 routes to somatic
embryogenesis.
• 1. Direct embryogenesis: The embryo initiates
directly from the explant tissue in absence of
callus proliferation.
• This occurs through pre-embryonic determined
cells (PEDC). Such cells are found in embryonic
tissues.
• Eg: scutellum of cereals
• 2.Indirect embryogenesis: The embryos initiate
from the explant through callus proliferation.
• This occurs through Induced embryogenic
determined cells (IEDC).
• Eg: Secondary phloem of carrot, leaf tissue of coffee,
petunia, asparagus etc.
• For some species, any part of plant body serves as an
explant, for embryogenesis. But in some species only
certain regions of explant may respond in culture. Eg:
cereals
• Floral or reproductive tissue in general has proven to
be an excellent source of embryogenic material.
• The physiological states of the plant from which
explant is taken is also extremely important.
STAGES OF SOMATIC EMBRYOGENESIS
• Somatic embryogenesis encompasses
various stages such as
• 1. Callus initiation
• 2. Embryo development and maturation
• 3. Plantlet formation
Endosperm culture and somatic embryogenesis
FACTORS AFFECTING SOMATIC
EMBRYOGENESIS
• 1. Genotype of explants: Explant genotype
has a marked influence on somatic embryo
regeneration and in many cases, it may
determine whether or not somatic embryo
regeneration will occur.
• Strong genotypic effect have been shown in
many species.
• Eg: Alfa-alfa , wheat, maize, rice and chick pea
etc.
• 2. Growth regulators:
• Auxins: Auxin alone or in combination with
cytokinin appear essential for the onset of growth
and the induction of embryogenesis of all the auxins,
2,4-D followed by NAA have proven to be extremely
useful.
• Effective concentration ranges are 0.5 – 27.6 μ M for
2,4-D and 0.5 – 10.7 μ M for NAA.
• Cytokinins: CKs have been used in the primary
medium invariably during embryogenesis of crop
plants.
• Effective concentrations for kinetin0.5 – 50.0μ M.
CKs are important in fastening somatic embryo
maturation and especially cotyledon development.
• ABA: It is added at inhibitory levels @0.1- 1 μM
promotes somatic embryo development and
maturation and at same time inhibits abnormal
proliferation and initiation of accessory embryos.
• 3. Nitrogen source: Form of nitrogen has marked
influenced on somatic embryogenesis. In carrot NH₄⁺
form has a promotive effect.
• Somatic embryos development occurs on a medium
containing NO₃⁻ as the sole nitrogen source.
• 4. Other factors: High K⁺ levels and low dissolved
O₂ levels prevents somatic embryo regeneration.
• In citrus medica, volatile compounds like
ethanol, inhibit somatic embryo regeneration.
• In soybean low sucrose conc. @ 5 to 10 g/l
promotes somatic embryogenesis than high
concentration.
• In Alfa-alfa, use of maltose as a carbon source
improved both somatic embryogenesis
induction and maturation as compared to
sucrose.
SYNTHETIC SEED CONCEPT
• Synthetic seeds are also known as artificial
seeds and also encapsulated embryos.
• The aim of somatic embryo encapsulation is to
produce an analog to true seeds, which is
based on the similarity of somatic embryo
with zygotic embryo with respect to their
morphology, physiology and biochemistry.
• Two type of synthetic seeds have been
developed namely,
• 1. Hydrated S.S.
• 2. Dessicated S.S.
• 1. Hydrated synthetic seeds: Renderbergh et al
(1986) developed hydrated artificial seeds by mixing
somatic embryos with an encapsulation matrix so
that embryos are well protected in that matrix and it
is rigid enough to allow for rough handling.
• Encapsulation matrix consists of hydrogels such as
agar, carrageenan, alginate or plant exudates of
arabica, karaya or seed gums of guar, tamarind or
microbial products like dextran, xantham gum and
divalent salts.
• Various divalent salts which can be used are CaCl₂,
Ca(OH)₂, ferrous chloride, cobaltous chloride etc.
• Sodium alginate + CaCl₂ = encapsulation of somatic
embryo.
• Procedure: Mixing the somatic embryos with
sodium alginate, followed by dropping into a solution
of CaCl₂, CaNO₃ to form calcium alginate beads.
• Calcium alginate capsules tend to stick together and
are difficult to handle because they lose water
rapidly and dry down to a hard pellet within a few
hours of exposure to the atmosphere.
• These problems can be offset by coating capsules
with Elvax 4260 (Ethylene vinyl acetate acrylic acid
tetrapolymer)
• Sodium alginate + CaCl₂= Calcium alginate
• 2. Dessicated synthetic seeds: Kim and Janic
(1989)applied synthetic seed coats to clumps
of carrot somatic embryo to develop
dessicated artificial seeds.
• Mixing of equal volume of embryo suspension
and 5% of polyethylene oxide (a water soluble
resin), which subsequently dried to form
polyembryonic dessicated seeds.
• The survival of encapsulated embryos was
further achieved by embryo hardening
treatments with 12% sucrose or 10⁻⁶ M ABA,
followed by chilling at high inoculum density.
• Potential uses of synthetic seeds:
• Produce large number of identical embryos .
• Determines the roles of endosperm in embryo
development and germination.
• Propagation with low cost, high volume
capabilities of seed propagation.
• Produced within short time and in any season,
at any time.
• Dormancy period can be reduced there by
shortening the life cycle of a plant.
• Useful in preserving germplasm.
• Provides knowledge to understand the
developmental, anatomical characteristics of
endosperm and seed coat formation.
• Synthetic seeds production has been
successfully obtained in Zea mays, Opium,
Daucus carota, Lactuca sativa, Madicago,
Brassica species, Gossypium, Santalum species
etc.
• Disadvantages with artificial seeds:
• 1. Facilities required are costly.
• 2. Special skills are required.
• 3. Errors in maintenance.
• 4. Specific kinds of genetic and epigenetic
modifications can develop.
• Problems in production of synthetic seeds:
• 1. Artificial seeds that are stable for several months
requires the procedures for making embryos
quiscent.
• 2. Synthetic seeds need to be protected
against dessication.
• 3. Recovery of plants is often very low due to
incomplete embryo formation or difficulties in
creating artificial endosperm.
• 4. Embryo must be protected against
microbes.
Endosperm culture and somatic embryogenesis

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Endosperm culture and somatic embryogenesis

  • 2. • TOPIC: • TRIPLOID PRODUCTION (ENDOSPERM CULTURE)- DIFFERENTIATION AND FACTORS AFFECTING THEM- APPLICATIONS • SOMATIC EMBRYOGENESIS- FACTORS AFFECTING, STAGES OF SOMATIC EMBRYOGENESIS, LARGE SCALE PRODUCTION, SYNTHETIC SEEDS CONCEPT AND ITS PRO’S AND CON’S PRESENTED BY ZUBY GOHAR ANSARI TAM/14-26
  • 3. ENDOSPERM CULTURE (HISTORY AND DEF.) • Double fertilization occurs in majority of angiosperms which results in two fusion products i.e. Zygote (fusion product of egg and one of the male gamete) and triploid endosperm (fusion product of polar nuclei and second male gamete). • The endosperm lack many organogenic potential and vascular differentiation. • Endosperm is unique tissue, firstly in its function of supplying nutrition to developing embryo and secondly in being triploid.
  • 4. • Attempts to grow endosperm tissue in cultures began in 1930’s and now mature and immature endosperm of various taxa of angiosperm have been grown. • Lampe and Mills (1933) : They were first to report the proliferation of immature endosperm tissue of maize grown on medium containing extract of potato. • Nakano et al. (1975): Successfully cultured immature endosperm of rice and achieved organogenesis.
  • 5. • Rangaswami and Rao (1963): Successfully cultured mature endosperm of santalum album (chandan) up to callus. • Endosperm culture: It is the in vitro development of isolated mature or immature endosperm from seed at proper stage on a suitable culture medium to obtain triploid plantlet.
  • 7. TYPES OF ENDOSPERM CULTURE • 1. Mature endosperm culture: It is the in vitro development of isolated mature endosperm from ripen endospermic seed on suitable culture medium to obtain triploid plantlet is called mature endosperm culture. • 2. Immature endosperm culture: It is the in vitro development of isolated immature endosperm isolated at precise stage from immature seed, cultured on suitable culture medium to obtain triploid plantlet is called immature endosperm culture.
  • 8. STEPS IN ENDOSPERM CULTURE • It consist of mainly 3 steps: • 1. The immature or mature seeds are dissected under aseptic conditions and endosperms along with embryos are excised. • 2. Excised endosperms are cultured on a suitable medium and embryos are removed after initial stage. • 3. Callus followed by embryogenesis or shoot bud differentiation. • 4. Complete plant formation.
  • 10. PROCEDURE OF ENDOSPERM CULTURE • 1. Explant source: In most of the cereals, mature endosperm does not respond to cultural conditions. It is therefore, excised at proper period of development. Normally the endosperm of cereals undergo certain changes 12 days after pollination making this able to respond. Proper stage for maize is 8-11 days after pollination, 4-7 days after pollination for rice, and 8 days after pollination for wheat. • In some families association of embryo tissue in initial stages seems essential for proliferation of mature endosperm. In such cases, entire seeds are used as explant. Immature seeds provide explant for non- endospermic seeds.
  • 11. • 2. Inoculation of explant: For in vitro culture of mature endosperm, seeds having massive endosperm are decoated, surface sterilized with suitable disinfectant and after 2-3 washings with sterile distilled water, planted on the nutrient medium. For in vitro culture of an immature endosperm, the entire seed or kernel is surface sterilized and the endosperm tissue is excised under aseptic conditions. In cereals, top of kernel or immature ovaries (micropyle end) are cut off with a sterile knife and exposed endosperm squeezed out and placed on the callus induction nutrient medium. • The basal white and MS medium is generally used to induce callus from an immature endosperm.
  • 13. • The basal medium is supplemented with tomato juice, yeast extract, grape juice. Sucrose (2-4%) is used as a source of carbohydrates. In some cases, addition of 2,4- D or IAA, cytokinin necessary. • 3. Incubation/ maintenance of culture: To induce callus, the endosperm cultures are maintained darkness or diffused light. Differentiation take place when calli are transferred to bright light (2000 lux – 4000 lux) and 25⁰C temperature
  • 14. • 4. Shoot bud differentiation or embryogenesis: Plantlet formation from endosperm callus maturation follow organogenic or embryogenic mode of development. Embryo differentiation occurs when the proliferated tissue is transferred from callusing to basal medium with or without gibberellins. So culturing carried out up to complete development of plantlet. • 5. plantlet developed are hardened in green house by transferring in vermiculture media and maintaining proper humidity.
  • 15. FACTORS AFFECTING ENDOSPERM CULTURE • 1. Explant stage: proper stage may vary from cellular (immature) to mature endosperm depending upon species. (4-7 DAP in rice, 8 DAP in wheat and 12 DAP in maize and fully matured in santalum album). • 2. Nutrient medium: Low amount of reduced nitrogen is required for proliferation. Undefined source like tomato juice, yeast extract, etc.
  • 16. • 3. Physical factors: The pH 7.0 seems to be effective for fresh weight increase. Maximum growth of endosperm occurs between 24- 27⁰C temperature and 12-16 hours photoperiod with diffuse day light supported callusing as well as regeneration. • 4. Embryo factors: association of embryo tissue in the initial stages seems essential for inducing proliferation of mature endosperm tissue in some families. Immature seeds of non endospermic seeds exhibit no dependence on embryo factor.
  • 17. APPLICATIONS OF ENDOSPERM CULTURE • Techniques of endosperm culture has enabled the production of triploid plants. Triploid plants are self sterile and usually seedless. The trait increases edibility of fruits and is desirable in plants such as apple, banana, grape, watermelon and mango which are commercially important. • In timber and fuel yielding plants, triploids show better performance over their relative diploids or tetraploids. Also there is no problem of seed sterility as they can be multiplied by vegetative means.
  • 18. SOMATIC EMBRYOGENESIS • Regeneration of embryos from somatic cells, tissues or organs either denovo or directly in vitro conditions is known as somatic embryogenesis. • It is also known as non- zygotic/ non-sexual embryogenesis. • Various terms for non-zygotic embryos have been reported such as, • A. Adventive embryos: Somatic embryos arising directly from other organs or embryos.
  • 19. • B. Parthenogenic embryos: Formed by unfertilized eggs. • C. Androgenetic embryos: Formed by male gametophyte. • Occurence of asexual embryogenesis is generally restricted to intraocular tissues.
  • 21. • Differences between somatic and sexual embryos SOMATIC EMBRYOS SEXUAL EMBRYOS Embryos arises from single cell. Arises from multi cell. Embryos have bipolar structure. It is monopolar structure. Embryos has no vascular connections with cultured explant. Embryos has vascular connections with cultured explant. Induction of somatic embryogenesis requires single hormonal signal. Requires two hormonal signals.
  • 22. • The initiation and development of embryos from somatic tissues in plant culture was first recognized by steward et al (1958) and Reinert (1958- 59) in cultures of Daucus carota (carrot). • In addition to the development of somatic embryos from sporophytic cells, embryos have been obtained from generative cells, such as the classic works done by Guha and Maheswari (1964), with Datura innoxia microspores.
  • 23. TWO ROUTES TO SOMATIC EMBRYOGENESIS • Sharp et al (1980) divided 2 routes to somatic embryogenesis. • 1. Direct embryogenesis: The embryo initiates directly from the explant tissue in absence of callus proliferation. • This occurs through pre-embryonic determined cells (PEDC). Such cells are found in embryonic tissues. • Eg: scutellum of cereals
  • 24. • 2.Indirect embryogenesis: The embryos initiate from the explant through callus proliferation. • This occurs through Induced embryogenic determined cells (IEDC). • Eg: Secondary phloem of carrot, leaf tissue of coffee, petunia, asparagus etc. • For some species, any part of plant body serves as an explant, for embryogenesis. But in some species only certain regions of explant may respond in culture. Eg: cereals • Floral or reproductive tissue in general has proven to be an excellent source of embryogenic material. • The physiological states of the plant from which explant is taken is also extremely important.
  • 25. STAGES OF SOMATIC EMBRYOGENESIS • Somatic embryogenesis encompasses various stages such as • 1. Callus initiation • 2. Embryo development and maturation • 3. Plantlet formation
  • 27. FACTORS AFFECTING SOMATIC EMBRYOGENESIS • 1. Genotype of explants: Explant genotype has a marked influence on somatic embryo regeneration and in many cases, it may determine whether or not somatic embryo regeneration will occur. • Strong genotypic effect have been shown in many species. • Eg: Alfa-alfa , wheat, maize, rice and chick pea etc.
  • 28. • 2. Growth regulators: • Auxins: Auxin alone or in combination with cytokinin appear essential for the onset of growth and the induction of embryogenesis of all the auxins, 2,4-D followed by NAA have proven to be extremely useful. • Effective concentration ranges are 0.5 – 27.6 μ M for 2,4-D and 0.5 – 10.7 μ M for NAA. • Cytokinins: CKs have been used in the primary medium invariably during embryogenesis of crop plants. • Effective concentrations for kinetin0.5 – 50.0μ M. CKs are important in fastening somatic embryo maturation and especially cotyledon development.
  • 29. • ABA: It is added at inhibitory levels @0.1- 1 μM promotes somatic embryo development and maturation and at same time inhibits abnormal proliferation and initiation of accessory embryos. • 3. Nitrogen source: Form of nitrogen has marked influenced on somatic embryogenesis. In carrot NH₄⁺ form has a promotive effect. • Somatic embryos development occurs on a medium containing NO₃⁻ as the sole nitrogen source. • 4. Other factors: High K⁺ levels and low dissolved O₂ levels prevents somatic embryo regeneration.
  • 30. • In citrus medica, volatile compounds like ethanol, inhibit somatic embryo regeneration. • In soybean low sucrose conc. @ 5 to 10 g/l promotes somatic embryogenesis than high concentration. • In Alfa-alfa, use of maltose as a carbon source improved both somatic embryogenesis induction and maturation as compared to sucrose.
  • 31. SYNTHETIC SEED CONCEPT • Synthetic seeds are also known as artificial seeds and also encapsulated embryos. • The aim of somatic embryo encapsulation is to produce an analog to true seeds, which is based on the similarity of somatic embryo with zygotic embryo with respect to their morphology, physiology and biochemistry. • Two type of synthetic seeds have been developed namely, • 1. Hydrated S.S. • 2. Dessicated S.S.
  • 32. • 1. Hydrated synthetic seeds: Renderbergh et al (1986) developed hydrated artificial seeds by mixing somatic embryos with an encapsulation matrix so that embryos are well protected in that matrix and it is rigid enough to allow for rough handling. • Encapsulation matrix consists of hydrogels such as agar, carrageenan, alginate or plant exudates of arabica, karaya or seed gums of guar, tamarind or microbial products like dextran, xantham gum and divalent salts. • Various divalent salts which can be used are CaCl₂, Ca(OH)₂, ferrous chloride, cobaltous chloride etc. • Sodium alginate + CaCl₂ = encapsulation of somatic embryo.
  • 33. • Procedure: Mixing the somatic embryos with sodium alginate, followed by dropping into a solution of CaCl₂, CaNO₃ to form calcium alginate beads. • Calcium alginate capsules tend to stick together and are difficult to handle because they lose water rapidly and dry down to a hard pellet within a few hours of exposure to the atmosphere. • These problems can be offset by coating capsules with Elvax 4260 (Ethylene vinyl acetate acrylic acid tetrapolymer) • Sodium alginate + CaCl₂= Calcium alginate
  • 34. • 2. Dessicated synthetic seeds: Kim and Janic (1989)applied synthetic seed coats to clumps of carrot somatic embryo to develop dessicated artificial seeds. • Mixing of equal volume of embryo suspension and 5% of polyethylene oxide (a water soluble resin), which subsequently dried to form polyembryonic dessicated seeds. • The survival of encapsulated embryos was further achieved by embryo hardening treatments with 12% sucrose or 10⁻⁶ M ABA, followed by chilling at high inoculum density.
  • 35. • Potential uses of synthetic seeds: • Produce large number of identical embryos . • Determines the roles of endosperm in embryo development and germination. • Propagation with low cost, high volume capabilities of seed propagation. • Produced within short time and in any season, at any time. • Dormancy period can be reduced there by shortening the life cycle of a plant. • Useful in preserving germplasm.
  • 36. • Provides knowledge to understand the developmental, anatomical characteristics of endosperm and seed coat formation. • Synthetic seeds production has been successfully obtained in Zea mays, Opium, Daucus carota, Lactuca sativa, Madicago, Brassica species, Gossypium, Santalum species etc.
  • 37. • Disadvantages with artificial seeds: • 1. Facilities required are costly. • 2. Special skills are required. • 3. Errors in maintenance. • 4. Specific kinds of genetic and epigenetic modifications can develop. • Problems in production of synthetic seeds: • 1. Artificial seeds that are stable for several months requires the procedures for making embryos quiscent.
  • 38. • 2. Synthetic seeds need to be protected against dessication. • 3. Recovery of plants is often very low due to incomplete embryo formation or difficulties in creating artificial endosperm. • 4. Embryo must be protected against microbes.

Notes de l'éditeur