Contents

1. Concept of Plant Tissue Culture

1. Concepts of Cell Theory

2. Cellular Totipotency

3. Landmarks in Plant Tissue Culture

4. The General Technique

4.1 Important Features of Tissue Culture Technique

4.2 Importance of Tissue Culture Technique

5. Applications of Plant Tissue Culture

2. Basic Facilities of Plant Tissue Culture

1. General and Aseptic Laboratory

2. Different Work Areas

2.1 Culture Room

2.2 Bottle Wash Machine

2.3 Hand and Leg Wash Room

2.4 Dressing Room

2.5 Culture Transfer Room

3. Equipments and Instruments used in Plant Tissue Culture

4. Laboratory – Research Lab and Commercial Lab

4.1 Research Laboratory for Plant Tissue Culture

4.2 Commercial Laboratory for Plant Tissue Culture

3. Aseptic Techniques

1. Introduction

2. Washing and Preparation of Glassware

3. Packing and Sterilization

3.1 Packing

3.2 Sterilization

4. Aseptic Work Station

5. Precautions to maintain Aseptic Conditions

4. Media Preparation

1. Composition of M. S. Medium

2. General Methodology for Medium Preparation

3. Different Types of Media used in PTC

4. Plant Growth Regulators (PGR's) and their In Vitro Roles

5. Techniques of Plant Tissue Culture

1. Introduction

2. Root Tip Cultures

2.1 Factor affecting Root Tip Culture

3. Leaf culture

3.1 Factor affecting Leaf Culture

4. Shoot Tip Culture

5. Meristem Culture

5.1 Meristem Culture

5.2 Media Preparation

5.3 Application of Meristem Culture

6. Importance of Organ Culture

6. Micropropagation

1. Concept of Differentiation

2. Types of Micropropogation

3. Stages of Micropropagation

4. The Various Applications of Micropropagation

7. Callus Culture Techniques

1. Introduction

2. Principle

3. Callus Cultures

3.1 Protocol for the Culture of Plant Tissues

3.2  Significance of Callus Culture

3.3 Morphology

3.4 Factors Affecting Callus Culture

3.5 Internal Structure of Callus

3.6 Maintenance of Callus

3.6 Application and Limitation of Callus

8. Suspension Culture Techniques

1. Introduction

2. Types of Suspension Cultures

3. Culture Medium

4. Synchronization

5. Protocol for the Establishment of Suspension Culture

6. Assessment of Growth

6.1 Cell Packing Method

6.2 Direct Cell Count Method

6.3 Direct Weight Method

6.4 Protein Measurement Technique

7. Experiments to assess the Cell Viability

7.1 Evan's Blue Dye Treatment

7.2 Chemical Method

8. Importance and Uses of Suspension Culture

9. Organogenesis

1. Introduction

2. Protocol

3. Types of Organogenesis

4. Factors Affecting Organogenesis

5. Applications and Limitations of Organogenesis

5.1 Applications

5.2 Limitations

10. Somatic Embryogenesis

1. Introduction

2. Types of Embryogenesis

3. Stages of Somatic Embryogenesis

4. Factors Affecting Somatic Embryogenesis

5. Applications of Somatic Embryogenesis

6. Limitations of Somatic Embryogenesis

7. Production of Artificial Seeds

7.1 Techniques to Produce Artificial Seeds

7.2  Types of Artificial Seeds

7.3 Salient Features of Artificial Seeds

11. Anther and Pollen Culture

1.  Anther Culture

1.1  Protocol for Pollen Culture Techniques

1.2  Principle

1.3  Factors Affecting Anther Culture

2. Pollen Culture

2.1  Protocols for Pollen Culture Technique

2.2  Principle

2.3  Factors Affecting Pollen Culture

2.4 Comparison between Anther and Pollen Culture

2.5 Pollen Biotechnology

12. Ovary, Ovule, Embryo and Endosperm Culture

1. Introduction

2.  Ovary Culture

3. Ovule Culture

4. Embryo Culture

4.1 Culturing Method

4.2 Embryo Rescue

5. Endosperm

6. Factors Affecting Ovary, Ovule, Embry and Endosperm Culture

7. Applications and Limitations

7.1 Applications

7.2 Limitations

13. Protoplast – Isolation, Culture and Fusion

1. Methods of Isolation of Protoplasts

1.1 Mechanical Method

1.2  Enzymatic Isolation of Protoplasts

1.3 Two Step Enzymatic Isolation Method

2. Maintenance of Protoplasts

3. Culture of Protoplasts

3.1 Viability Tests for Protoplasts

3.2 Regeneration of Cell Wall

3.3 Cell Division and Callus Formation

3.4 Importance of Protoplast Culture

4. Somatic Hybridization

4.1  Protoplast Fusion

4.2 Selection of Hybrid Protoplasts

4.3  Regeneration of Plantlets

4.4 Production of Symmetric and Asymmetric Hybrids

4.5 Examples of Somatic Hybrids

4.6 Cybrids

5. Applications of Protoplast Fusion and Somatic Hybridization

6. Limitations of Protoplast Fusion and Somatic Hybridization

14. Genetic Transformations

1. Gene Manipulation in Plants

2. Direct DNA Transfer Methods

3. Transformation of Protoplasts

3.1 Methods for the Insertion of Organalle into the Protoplasts

3.2 Selection of Transformed Protoplast

3.3 Culture of Protoplasts

4. Agrobacterium Mediated Gene Transfer

4.1 Salient Features of Ti-plasmid

4.2 Types of Ti-Plasmids

4.3 Structure of T-DNA Fragment

4.4 Gene Transfer through Ti-Plasmids

4.5 Difficulties of Gene Transfer through Ti-Plasmid

4.6 Results of Gene Transfer Plants

5. Integration and Expression of Foreign DNA

5.1 Integration of Transgenes

5.2 Inheritance of Transgenes

5.3 Gene Expression

5.4 Gene Silencing

15. Applications of Plant Tissue Culture in Research and Commercial Industry

1. Applications of Cell and Tissue Culture

1.1 Micropropagation / Clonal Propagation

1.2 Production of Virus Free Plants

1.3 Production of Synthetic Seeds

1.4 Mutant Selection

1.5 Production of Secondary Metabolites

1.6 Somatic Hybridization

1.7 Cybrids

1.8 In Vitro Plant Germplasm Conservation

1.9 Transgenic Plants with Beneficial Traits

1.10 Delayed Fruit Ripening

1.11 Male Sterility

1.12 Transgenic Plants as Bioreactors (Molecular Farming)

1.13 Edible Vaccines

1.14 Biodegradable Plastics

1.15 Molecular Breeding

1.16 Commercial use of Transgenic Plants