Somatic Embryogenesis
Somatic Embryogenesis
Michael Greer
Somatic Embryogenesis
Somatic Embryogenesis
Potential to study early embryonic
development
– No integuments hiding development
Potentially very high yield
Single cell origin
Ploidy number alteration
Synthetic seed production?
Randall P. Niedz, Ph.DU. S. Department of Agriculture - Agricultural Research Service
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Somatic Embryogenesis
Potential to study early embryonic
development
Potentially very high yield
– 60,000 to 1 350 000 cells per litre
Single cell origin
Ploidy number alteration
Synthetic seed production?
Somatic Embryogenesis
Somatic Embryogenesis
Potential to study early embryonic
development
Potentially very high yield
Single cell origin
– Ideal for transformation techniques
I.e.. Microparticle bombardment
Ploidy number alteration
Synthetic seed production?
Somatic Embryogenesis
Potential to study early embryonic
development
Potentially very high yield
Single cell origin
Ploidy number alteration
Synthetic seed production?
www.bsbanet.org/imi/dolgov1/image002.jpg
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Jimenez (2001)
In order to better understand and recreate
somatic embryogenesis, Jimenez
assayed the endogenous hormone levels
of both initial explants and the produced
calli from highly regenerable and
recalcitrant lines.
The usual suspects
Cytokinins
– BAP, kinetin, zeatin,
zeatin, 2-iP
– Can initiate somatic embryogenesis in some
species
– Often used for secondary embryogenesis
The usual suspects
Auxin
– 2,4-D, NAA, IAA, IBA etc.
– Induces change in gene expression to initiate
development of globular embryo
Initiator of somatic embryos
Must be reduced to establish polar auxin
gradient
The usual suspects
Abcisic acid
– ABA
– Seed formation, stress response
– Peaks during embryo maturation
– Lowest as seed desiccation
– Necessary for monocot embryo initiation
– Regulate carbohydrate metabolism
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The usual suspects
Jimenez (2001)
endogenous explants hormone levels
Gibberellins
– GA3, GA1, GA20
– Elongation of internodes
– Inhibit shoot formation and adventitious root
– Necessary at all for somatic embryogenesis?
Jimenez (2001)
Jimenez (2001)
endogenous calli hormone levels
Numerous conflicting reports
Seems that every tissue has a sensitivity
to the different hormones which is equal
to or more important then the actual
concentration
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Madden (et al. 2005) found
that in flowers
(Pelargonium x Hortorum)
Hortorum)
that the embryos
produced possessed all
the feature of somatic
embryos
Madden (et al. 2005) found
that in flowers
(Pelargonium x Hortorum)
Hortorum)
that the embryos
produced possessed all
the feature of somatic
embryos,
But these somatic
embryogenesis seemed
to lack the formation of a
root pole.
Bassuner (et al. 2006)
Before we can move on and improve
somatic embryogenesis protocols in nonmodel systems, we must first understand
whether or not we are in fact inducing
somatic embryogenesis in the first place.
Bassuner (et al. 2006) generated a
PIN4/LENNY enhancer trap line with GUS
as a reporter gene.
AtPIN4 is a auxin efflux protein which
involved in channelling the hormone to
create a ‘auxin sink’
sink’ which is necessary
for proper development of the root.
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Bassuner (et al. 2006)
Zygotic PIN4 expression
Dr5::GUS was also employed to elucidate
auxin response
Zygotic PIN4 expression
Embryonic PIN4 expression
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Embryonic PIN4 expression
Embryonic PIN4 expression
J.& K. Adventitious shoot, L. Fused shoot, M. Adventitious shoot
Embryonic DR5::Gus expression
A. Zygotic embryo
B. Somatic embryo
C. Adventitious shoot
D. Somatic embryo
E. Adventitious shoot
Three main regenerative structures
– 1. adventitious shoots
– 2. fused shoots
– 3. true somatic embryos
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Liquid versus Solid Medium
Nhut (et al. 2006)
Three main regenerative structures
– 1. adventitious shoots
– 2. fused shoots
– 3. true somatic embryos
True somatic embryos require proper auxin
transport to develop
Liquid versus Solid Medium
Nhut (et al. 2006)
Somatic Embryogenesis
Thank You
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Refferences
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gradients establish the apical-basal axis of Arabidopsis Nature. 2003 Nov 13;426(6963):147-53
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Lukowitz,, W.; Yeung
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anh, N. T. M.; Tuan, P. Q.; Nguyet
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embryogenesis of Lilium longiflorum Scientia Horticulturea 110:93-97
Jim enez, V. M.; Bangerth (2001) Horm onal status of m aize initial explants and of the em bryogenic and nonem bryogenis callus cultures derived from them as related to m orphogenesis in vitro plant science 160:247-257
Shultz, E. (2006) Plant development course lectures (5560) http://classes.uleth.ca/200603/biol4560a
Chawla,, H. S. (2002) Introduction to plant biotechnology second edition Science publishers,
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usa
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