For B.Pharm, 4th Sem

1. Somatic
hybridization

2. Definition
• Fusion of protoplasts facilitates mixing of
two whole genomes and could be
exploited in crosses at inter-specific,
inter-generic, or even inter-kingdom
levels.
• Isolated protoplasts are devoid of cell
walls make them easy tools for
undergoing fusion in vitro.

7. Cybrid
• Plants or cells which containing nucleus of one
species but cytoplasm from both the parents
species.
• Its a cyto-plasmic hybrid
• Involved fusion of two protoplasts (donor &
recipient) in which donor nuclear genome is
eliminated while its plastomes are merged
with that of the recipient.
Suppression of nuclear genome during
fusion(uses gama rays & x-rays

8. Somatic hybridization techniques

11. 1. Spontaneous fusion
• Protoplasts fuse spontaneously
during isolation process mainly
due to physical contact.
(physical contact is sufficient)
• It is strictly Intra-specific &
produce homokaryons.

12. 2. Induced fusion
• Normally, isolated protoplasts do not fuse with
each other easily because the surface of the
isolated protoplast carries negative charge( -10mV
to -30mV)
• Protoplast from widely different and sexually
incompatible plants can be fused by this
procedure
• Classified into three groups-
i) Chemofusion-fusion induced by chemicals
ii) Mechanical- fusion induced by mechanically
iii) Electrofusion- fusion induced by electrical
stimulation

13. i) Chemofusion-fusion induced by
chemicals
• Examples of Chemical fusogens
1. PEG
2. NaNo3
3. Ca 2+ ions
4. Polyvinyl alcohol
• Agents which are commonly known as chemical
fusogens
• Chemical fusogens cause the isolated
protoplasts to adhere to one another and leads to
tight agglutination followed by fusion of
protoplast.

14. ii) Mechanical fusion
• Protoplasts are brought into intimate physical
contact mechanical under microscope using
micro-manipulator and perfusion
micropipette.
• Micro-pipette is partially blocked within 1mm
of the tip by a sealed glass rod result
protoplasts are retained and compressed by
the flow of liquid.
• This will facilities the mechanical fusion of
protoplasts.

16. iii) Electrofusion
• Electrical stimulation is being used to fuse
protoplasts-method called electro-fusion of
protoplasts
• Protocol: two glass capillary microelectrodes are
placed in contact with the protoplasts. An electric
potential (10kv/m) give rise to di-electrophoretic
dipole generation within the protoplast suspension
leads to pearl chain arrangement of protoplast,
application of high intensity of electric impulse
(100kv/m) results in the electric breakdown of
memberene and subsequent fusion.

18. ASSGNMENT: Chemofusion
• Fusion by Sodium/Potassium Nitrate
• Fusion by Calcium ions at high pH
• Fusion by PEG

19. Applications of somatic hybridization
• Production of novel inter-specific and inter-
generic hybrids. Pomato (Hybrid of potato and
tomato).
• Production of fertile diploids and polyploidy
from sexually sterile haploids and triploids.
• Transfer gene for disease resistance, a biotic
stress resistance, herbicide resistance and many
other quality characters.

20. • Production of heterozygous lines in the single
species which cannot be propagated by
vegetative means.
• Formation of cytoplasm containing organelles
of two different parents.

23. Pomato plant

24. Limitation
• Poor regeneration of hybrid plants.
• Non-viability of fused products.
• Not successful in all plants.
• Production of un-favorable hybrids.
• Lack of an efficient method for selection of
hybrids.
• No confirmation of expression of particular
trait in somatic hybrids.