RAJESH BHAMMAR

y PLANT TISSUE CULTURE y PRINCIPLE

A. INTRODUCTION B. PLASTICITY C. TOTIPOTENCY D. CYTO-DIFFERENTIATION E. GENETICS F. PROCESS OF PLANT TISSUE CULTURE y APPLICATIONS IN A. STUDY B. AGRICULTURE C. MEDICINE D. PHARMA INDUSTRIES E. GENETICS F. R-DNA TECHNOLOGY
2

It is a technique of growing and maintaining the plant cell ,tissue or organ in a suitable medium in in vitro under aseptic condition.

Introduction

y Plants, due to their long life span, have developed a

greater ability to endure extreme conditions .

development adapt to environmental conditions.

growth and environment.

development

to

best

suit

their

y Particularly important aspects of this adaptation, as far as

plant tissue culture and regeneration are concerned, are the abilities to initiate cell division from almost any tissue of the plant y And to regenerate lost organs or undergo different developmental pathways in response to particular stimuli. y When plant cells and tissues are cultured in vitro they generally exhibit a very high degree of plasticity, which allows one type of tissue or organ to be initiated from another type. y In this way, whole plants can be subsequently regenerated.
6

y The ability of an individual cell to develop into a whole

plant is referred as to as cellular totipotency.

namely dedifferentiation and redifferentiation are responsible for the phenomenon of totipotency.

y The phenomenon of mature cell (differentiated)

reverting to meristematic state to produce callus is known as dedifferentiation.

y The ability of callus cells to differentiate into a plant

organ or a whole plant is regarded as redifferentiation.

10

11

12

y In plant tissue culture , during growth and maturation

of the callus tissue or free cells in suspension culture, few dedifferentiation cells undergoes cytoquiescence and cytosenescence.
y These two changes are mainly associated with the

process of cyto-differentiation which will ultimately form the vascular tissue , particularly xylem elements.
y This phenomenon is known as cytodifferentiation
13

14

As per principle

y Explant:
Any piece of the differentiated or undifferentiated tissue taken from the plant to start plant tissue culture. laboratory: 1. washing area 2. transfer area 3. culture room 4. green house Nutrient medium: Aseptic condition 1. incubator 2. autoclave 3 .lamina air flow
16

y y y

17

EXPLANT: y For callus culture may be the differentiated tissue from any part of the plant.
y Due to their genetic totipotency. y E.g. meristematic cell, leaf measophyll cell, phloem

cell, cambium Cells etc..

19

Cell suspension culture

21

y Undifferentiated and unorganized mass of plant cells. y Parenchymatous in nature although not truly

homogenous.
y Some quantity of differentiated tissue besides the bulk

of non-differentiated tissue.

22

y Explant and its genotype y Composition of medium y Physical factors y Age of the plant y Location of Explant y Growth regulators

23

y Callus is Slow growth plant culture system , this

enables to conduct several studies like 1. Nutritional requirement of plants 2. Cell and organ differentiation 3. Development of suspension and protoplast culture 4. Somaclonal variation 5. Product of secondary metabolites and their regulation

24

y Cell culture technique

Isolation of single cell 2. Callus culture 3. Suspension culture growth and subculturing 4. Synchronization of suspension culture
1.

25

To study pathways of cellular metabolism 2. Serve as good target for mutation and selection of desirable mutant 3. Production of secondary metabolites of commercial interest 4. Good potential for crop improvement
1.

26

Cell suspension culture

28

Introduction to the benefits

y Embryo culture y Ovary and ovule culture y Control of fertilization y Endosperm culture

30

y Anther culture y Somatic hybridization & genetic modification y Propagation through though tissue culture y Production of disease free plants y Synthetic seed production
31

y Protoplast from different species can be fused to

generate a hybrid and this process is known as somatic hybridization y New possibilities for the in vitro genetic manipulation of plant to improve the crops y Applications 1. Disease resistance 2. Environmental tolerance 3. Quality characters 4. Cytoplasmic male sterility 5. Study Cytoplasmic gene and their function
32

y The genetic variations found in the in vitro cultured cell are

collectively referred as Somaclonal variation. y Calliclones y protoclones Applications 1. Production of agronomically important useful plants. 2. Resistance to diseases 3. Resistance to abiotic stresses 4. Resistance to herbicides 5. Improve seed quality i.e. lathyrus seed.
33

CROP Rice Wheat Maize Sugarcane Tomato Carrot potato

CHARACTERS Flowering period; plant height; leaf length; shape and color; frequency of fertile seed Plant height; seed storage protein; grain color Reduced pollen fertility and male sterility High sugar yield; increase stalk length; diameter ;weight and density Early flowering; orange fruit color Higher carotene content Higher yield; growth habit; maturity and morphology
34

y High rate of plant propagation y Production of diseased free plant y Cost effective process y Automated micropropagation

35

y Prevention of embryo abortion y Overcoming seed dormancy y Shortening of breeding cycle y Production of haploids y Overcoming of seed sterility y Clonal propagation
36

y Possessing only a single set of chromosomes in the

sporophyte. y Applications: 1. Development of homozygous lines 2. Generation of exclusive male plant 3. Induction of mutation 4. Production of disease resistant plants 5. Production of insect resistant plant 6. Production of salt tolerance plant 7. Cytogenetic research 8. Induction of genetic variability 9. Evolutionary studies

37

y Increase in human population , pressure on the forest

and land resources have increased which result in decline in the population of medicinal and economically important plant species. y Threatened species y Conserve the Germplasm by preserving the genetic material, 1 . In situ preservation 2 . Ex situ preservation
38

y Most of the natural products are compound derived

y y y y

from primary metabolites such as amino acid, carbohydrate and fatty acids. Secondary metabolites are considered as the end product of primary metabolites. Plant tissue culture gives the excellent tool for study the various metabolic activity of the cell. From this study it is easy to produce medically important compounds (as secondary metabolite). E.g. Morphine, Codeine etc..
39

Ajmalicine

Digoxin

papain

reserpine

Atropine

Ephdrine

Papaverine

Scorpolanine

Caffeine

Hyoscyamine

Psedoephedrine

Sennosides

Codein

L-Dopa

Quinidine

Vincaleukoblastine

Colchicine

Morphine

Quinine

40

y Phytochemicals y Primary metabolites y Secondary metabolites y Application of secondary metabolites in 1. 2. 3. 4. 5. 6.

Pharmaceuticals Flavors Perfumes Agrochemicals Insecticides Raw material for industries

y Production under control conditions as per

demand y Culture system are independent of 1. environmental factors 2. seasonal variation 3. Pest and microbial diseases 4. Geographical constrains y Cell growth can be controlled to facilitate improved product formation y Quality of the product will be consistent as its product from specific cell line
42

y Recovery of product will be easy y Easy to provide desired environment than in the field y Mutant cell line can be developed y Biotransformation reactions can be carried out with

certain culture cells y The production time is less and labour cost are minimal

43

y The conversion of one chemical into another by using

biological system (cell suspension) . y Conversion of less important compound to valuable medicinal or commercially important product. y Bioconversion may involve many type of reaction e.g. Reduction etc y It is important to select such cell line that possess the enzyme for catalysing the desired reaction

44

Substrate Digitoxin Codeinone carvoxin vanilin steviol

Product Digoxin codeine cavaxone Vanillin-D-glucuside Steviocide;steviobiocide

45

y Expression and regulation of gene y Site directed mutagenesis y rDNA technology y Function of gene product

46

U.Satyanarayana,2007.callus culture, cell culture, secondary metabolites, somatic hybridization, production of haploid plant, Somaclonal variation, micropropagation, synthetic seeds, embryo culture, Germplasm conservation, 497-516,523-537, 538-545, 546-551, 552-564, 565-566, BIOTECHNOLOGY, Book and allied (p) Ltd. K.G.Ramawat, G.Shaily,2010.application of plant tissue culture in agriculture, medicine, horticulture, forestry, Germplasm preservation and industry.325340,molecular biology and biotechnology, S.chand
48

Kalyan Kumar de,1997,cytodifferentiation, totipotency, 55-58, 59-66, an introduction to plant tissue culture, new central book agency(p) Ltd. Important URL For videos: y http://www.youtube.com/watch?v=4EWju4JNcxc&p=7 AEB2041844CA224&playnext=1&index=6 y http://www.youtube.com/watch?v=oYkpu41LVgk y http://sciencehack.com/videos/view/jKMwLrbYyJ0 y http://www.youtube.com/watch?v=D7Glb3l9mbY y http://www.youtube.com/watch?v=Jew0j1RGgvM&feat ure=related
49

50

51