Tissue culture is the growth of tissue  or cells separate from the organism Strawberry (Fragaria x ananassa Duch.) is a natural hybrid of Fragaria chiloensis . It is a perinnial, stoloniferous herb belongs to the Rosaceae family. Strawberries have traditionally been a popular delicious fruit for its flavour, taste, fresh use, freezing and processing.

1. Plant tissue culture
Prepared by: Abedalru’of M. Abu Hawas
Jul 2,2017

2. Content
 Introduction
 Techniques Of Plant Tissue Culture
 Tissue culture of strawberry :
• Objective
• Material and Method
• Result
 Conclusion
 References

3. Introduction
 Tissue culture is the growth of tissue or cells separate from the organism, This is
typically facilitated via use of a liquid, semi-solid, or solid growth medium , such as broth
or agar.
 Tissue culture commonly refers to the culture of animal cells and tissues, with the more
specific term plant tissue culture being used for plants.

4. Introduction
 Plant tissue culture is a collection of techniques used to maintain or grow plant
cells, tissues or organs under sterile conditions on a nutrient culture medium of
known composition.
 Plant tissue culture is widely used to produce clones of a plant in a method
known as micropropagation .

5. Introduction
 Different techniques in plant tissue culture may offer certain advantages over
traditional methods of propagation, including:
• The production of exact copies of plants
• To quickly produce mature plants.
• The regeneration of whole plants from plant cells that have been genetically modified.
• The production of plants in sterile containers that allows them to be moved with greatly
reduced chances of transmitting diseases, pests, and pathogens.

6. Introduction
 Plant tissue culture relies on the fact that many plant cells have the ability
to regenerate a whole plant. Single cells, plant cells without cell walls
(protoplasts), pieces of leaves, stems or roots can often be used to
generate a new plant on culture media given the required nutrients
and plant hormones.

7. Techniques
 Preparation of plant tissue for tissue culture is performed under conditions
provided by a laminar flow cabinet .
 The tissue is grown in sterile containers, such as petri dishes or flasks in a growth
room with controlled temperature and light intensity.

8. Techniques
• Flasks containing tissue
culture growth medium
• Laminar flow cabinet

9. Techniques
 Living plant materials from the environment are naturally contaminated on their
surfaces with microorganism , so their surfaces are sterilized in chemical
solutions before suitable samples( known as explants ) are taken.
 The sterile explants are then usually placed on the surface of a sterile solid
culture medium.

10. Techniques
 Solid and liquid media are generally composed of inorganic salts plus a few
organic nutrients, vitamins and plant hormones .
 The composition of the medium, particularly the plant hormones and the
nitrogen source have profound effects on the morphology of the tissues that
grow from the initial explant .

11. Techniques
 As cultures grow, pieces are typically sliced off and subcultured onto new media
to allow for growth or to alter the morphology of the culture.
 The skill and experience of the tissue culturist are important in judging which
pieces to culture and which to discard.

12. Techniques
• Cultured cells growing in growth medium • strawberry plants produced by tissue
culture

13. Tissue culture of strawberry
Tissue culture of strawberryMicropropagation of Strawberry (Fragaria X ananassa
Duch)
S. Sakila, M.B. Ahmed, U.K. Roy, M.K. Biswas, 11 1 2 R. Karim, M.A. Razvy, M. Hossain, R. Islam and A.
Hoque

14. Tissue culture of strawberry
Objective :
•In the view of the potential commercial value, it is highly desirable to develop
methods of rapid, efficient and large scale multiplication of Fragaria X ananassa
Duch. through tissue culture.
•In the present study a simple protocol has been developed to propagate
strawberry through tissue culture methods from nodal segments in order to
ensure abundant supply of this plant material for commercial cultivation.

15. Tissue culture of strawberry
Tissue culture of strawberry
• Introduction :
Strawberry (Fragaria x ananassa Duch.) is a natural hybrid of Fragaria chiloensis
.
It is a perinnial, stoloniferous herb belongs to the Rosaceae family.
Strawberries have traditionally been a popular delicious fruit for its flavour, taste,
fresh use, freezing and processing.

16. Tissue culture of strawberry
Material and Method :
• Fresh nodes from strawberry mature plants were collected during the first week of
November 2006.
•Explants were washed in running tap water and then washed again thoroughly by
adding a few drops of Tween-20.

17. Tissue culture of strawberry
•They were then surface sterilized in a 0.1% mercuric chloride for 5 min followed by
rinsing them four times with double distilled water inside the Laminar Air flow
chamber.
•Small nodal segments (0.5–1.0 cm) were cultured on MS supplemented with specific
concentration of growth regulators (BA, KIN and GA ) singly or in combination adding
30 g l sugar (market sugar) and 0.7% agar.

18. Tissue culture of strawberry
•The pH of the medium was adjusted to 5.7 with 0.1 NaOH before autoclaving at 1.06
kg/cm and 121°C for 20 min.
•The cultures were incubated at 20 ± 2°C with 16 h photoperiod .

19. Tissue culture of strawberry
•Subcultures were done every 21 days interval
• Nodal segments from the proliferated shoots were subcultured again for further
multiple shoot induction.

20. Tissue culture of strawberry

21. Tissue culture of strawberry
• Regenerated multiple shoots were cut and individual shoots were placed in MS
medium containing different concentrations of IBA and IAA for root induction.
•Data were recorded after 5 weeks for recording multiple shoot induction and rooting
frequency.
•Only data which showed some advantageous effect were included in the tables and
25 explants were used per treatment and repeated three times.

22. RESULTS
•Nodal explants were inoculated on MS medium fortified with different
concentration of BA (0.5-2.0 mg l ) 1 w ith KIN (0.1 and 0.5 mg l ) or GA
(0.1and 0.5 mg l ).
•Within five weeks of culture multiple shoots emerged directly from the
explants.

23. RESULTS
•The numbers of shoots in medium with BA + KIN were greater than
those observed in the medium supplemented with BA + GA The highest
3. rate of response was obtained at 1.5 mg l BA+0.5 mg l 1 1 KIN
combination where 88% explants showed shoot proliferation and
9.3±058 shoots developed.

24. RESULTS

25. RESULTS
•When BA concentration was increased above 1.5 mg l , the rate 1 of
shoot multiplication reduced.
•However, the maximum number of shoots per explant and highest
average length were recorded at 1.5 mg l BA+0.1 mg l KIN. When BA 1 1
was supplemented with GA instead of KIN the rate of 3 shoot
proliferation and number of shoot did not improve but the shoot length
increased.

26. RESULTS
•In the present study under the moderate combination of 1.5 mg l BA + 1
0.1-0.5 mg l KIN was found to be the ideal combination 1 for high
frequency multiple shoot induction.

27. RESULTS
• Shoot proliferation on MS supplemented with
1.5 mg/l BA+0.1 mg/l KIN after 7 days of
culture .

28. RESULTS
•Shoot proliferation on MS+1.5 mg/l BA+0.5
mh/l KIN after 20 days of culture .

29. RESULTS
•Shoot proliferation on MS+1.5 mg/l BA+0.5
mg/l KIN after 35 days.

30. RESULTS
•The developing shoots were elongated by subculturing on the same
combinations of growth regulators.
•Later on elongated shoots were excised and used for root induction.

31. RESULTS
• Out of different concentrations of IBA (0.1-1.5 mg l ) 1 and IAA (0.1–1.5
mg l ) tested 1.0 mg l IBA proved to 1 1 be the most suitable for root
induction with 5.0±0.23 roots per explant and the average root length
being 3.68 cm

32. RESULTS

33. RESULTS
•Rooted shoots on 1.0 mg/l IBA
after 3 weeks of culture .

34. RESULTS
•Rooted plantlets were taken out from culture tubes and washed
thoroughly with tap water to remove the culture medium from the roots.
•Washed plantlets were sprayed with fungicide and planted to normal
and sterilized soil in polybags .

35. RESULTS
•Regenerated plantlets of
strawberry in polybags after 20
days of transplantation

36. RESULTS
• After 7 days the hardened plantlets were planted in soil.
•The protocol reported here is reproducible; it has a potential for allowing
a large scale micropropagation of this important and new plant .

37. References
• George, Edwin F.; Hall, Michael A.; De Klerk, Geert-Jan, eds. (2008). Plant
propagation by tissue culture
• S. Sakila, M.B. Ahmed, U.K. Roy, M.K. Biswas, 11 1 2 R. Karim, M.A. Razvy,
M. Hossain, R. Islam and A. Hoque . (1994). Micropropagation of Strawberry
(Fragaria X ananassa Duch.) . American-Eurasian Journal of Scientific
Research 2 (2): 151-154, 2007

38. The End
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