Duration: 100 min Kultur Jaringan Agroteknologi TA 2021-2022 Universitas Siliwangi

REGENERASI IN VITRO
dan EMBRIOGENESIS
SOMATIK
Visi Tinta Manik, S.Si., M.Si.

Pertemuan ke-4 dan 5

OUTLINE :
1.

REGENERASI TANAMAN

2.

ORGANOGENESIS IN VITRO

3.

EMBRIOLOGI SOMATIK
REGENERATION
• Regenarasi -> menumbuhkan kembali bagian tubuh yang rusak atau
lepas/rrespon fisiologis pada hewan dan tanaman
• Regenerasi : repairing <-> formation of new organ
• Plants possess a high capacity to regenerate -> ‘totipotency’
❑ The meristems are responsible for
repair after injury
❑ A unique feature of plant
regeneration is the formation of
new organs from cut sites
7. Final wrap-up ❑ In addition to these apical meristems,
plant stems also display several types
of tissue regeneration, including
vascular reformation after debarking
(Stobbe et al., 2002)
cytodiffrentiation
 • A common mode of plant regeneration is organogenesis
INDUKSI ORGANOGENESIS • Organogenesis adalah proses terbentuknya organ

Secara umum terdapat 3 fase organogenesis in vitro (Munetaka Sugiyama, 1999)

1. Sel pada eksplan mendapat kemampuan ‘kompeten’ (acquiring competence)
2. sel-sel kompeten dirangsang untuk membentuk organ tertentu dibawah pengaruh
keseimbangan hormon
3. Morfogenesis berlangsung secara independen yang disuplai hormon eksogen

eksplan organ

dediferensiasi diferensiasi

7. Final wrap-up Sel kompeten Determinated cell

Induksii
INDUKSI ORGANOGENESIS
• Organogenesis in vitro ditentukan oleh rasio hormon eksternal (auksin dan sitokinin)
Organogenesis -> 1. langsung Eksplan -> Meristemoid -> Primordia
2. Tiidak langsung Eksplan -> Kalus -> Meristemoid -> Primordia
Oganogenesis Rule of thumb :
1. Auxin/cytokinin = 10:1 – 100-1 = indce root
2. Auxin/cytokinin = 1:1o – 1:100 = induce shoot
3. Auxyn/Cytoinin around 1:1 favors callus
growth

7. Final wrap-up AUXIN CYTOKININ

IAA TDZ
IBA BA (BAP)
NAA Kinetin
2,4-D 2-ip
2,4,6,- T Zeatin
Dicamba
Picloram
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Faktor yang mempengaruhi diferensiasi tunas/pucuk
Exeption of the rule AxC :
1. Endogenous level of two GR

2. The level of cell sensitivity to GR

3. Uptake of GR
1. Kultur Media : konsep rasio auksin x 4. Degre og glycoslation and hydrolisis of GR

sitokinin 5. The type of GR

6. The activity of auxin and cytokinin oxidase

2. Genotip : regenerative potential ( high,
intermediete, less )
3. Explant : the source, the age, the plant
from which it is derived, preparation,
the manner it is planted on the medium.
4. Electrical and ultrasound stimulation of
shoot differentiation
EMBRIOGENESIS SOMATIK
TERMINOLOGI

Embryo : a new individual derived from single cell with defined shoot and root apex
having no vascular connection with maternal tissue

Zygotic embryogenesis : formation of embryo from zygotes

Non-zygotic embryogenesis : formation of embryo from cell other than zygotes

- Parthenogenetic embryogenesis
- Androgenetic embryogenesis
- Somatic embryogenesis

9
10
SOMATIC EMBRIOGNESIS VS ZIGOTIK EMBRIOGENESIS

(Zimmerman, 1993)
Fase Globular Fase Heart Fase Torpedo Fase Kotiledon
INDUKSI SE -> Langsung dan tidak langsung

(Tian, et al., 2020)

Eksplan :
1. Memiliki sifat Pre-embrionic Determined Cells (PEDCs) -> Direct SE

2. Induced Embrionic Determinated Cells (IEDCs) -> Indirect SE

3. Secondary embriogenesis (repetitive embriogenesis) can occurs

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INDUKSI SE Faktor yang mempengaruhi keberhasailan induksi SE :

min

1. Tahapan lengkap SE : induksi

embriogenesis – 1. Eksplan : limited to immature or less differentiated tissues
perkembangan embrio – 2. Genotip
pendewasaan embrio – 3. Medium : type of C sources, type of N source
4. GR : most commonly used is 2,4-D / zeatin
konversi/germinasi
5. Selective subcultur : embryonic callus vs non
2. Metode umum induksi SE
embryonic callus
dengan mengkultur eksplan
6. Electrical stimulation : affect the cell polarity
pada media sesuai dengan
penambahan auksin
7. Others : density, oxygen cons., prokaryot
3. Perkembangan setelah fase origin substanece (extract blue green algae),
globular membutuhkan light, etc
transfer ke media tanpa
auksin
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Tahapan kultur SE : min

1. Induksi : promote dedifferentiation (auxin), chamge cellular polarity

2. Development :
- cytoplasmicaly rich small cell -→ given rise to PEM (Proembryogenic Masses)/globules
- vacuolated long cells
- globules ->transfer-> globular
- synchronization SE development : Physical separation of different embrionic stage,
physiologically by GR (ABA)

3. Maturation/Conversion
- Promote maturation : ABA, High Sucrose, dessication
Keuntungan
1.Potensial untuk propagasi
skala besar (60.000 embrio/L
suspensi sel)
2.Adanya meristem pucuk dan
akar pada unit yang sama
3.Dormansi dan penyimpanan
jangka panjang
4.Kemungkinan enkapsulasi
(benih sintetis)
Penggunaan somatic embryogenesis dalam
crop improvement
1.Seleksi sel
2. Regenerasi tanaman yang
telah ditrasnformasi genetik
3. regenerasi hibrid somatik
4. produksi tanaman homozigt
dan poliploid
5.Preservasi germplasma
6. Eliminasi virus
7. produksi metabolit
8. teknologi benih sintetis
9.mikropropagasi
In woody plants, somatic embryogenesis plays a
critical role in clonal propagation and is a
powerful tool for synthetic seed production,
germplasm conservation, and cryopreservation
THANK YOU