Plant Tissue Culture Manual

Section A: Basic Techniques — Cell & Tissue Culture of Model Species.- 1. Media preparation.- 2. The initiation and maintenance of callus cultures of carrot & tobacco.- 3. The initiation and maintenance of plant cell suspension cultures.- 4. Shoot cultures and root cultures of tobacco.- 5. Somatic embryogenesis in orchardgrass.- 6. Arabidopsis regeneration and transformation (leaf & cotyledon expiant system).- 7. Arabidopsis protoplast transformation and regeneration.- 8. Arabidopsis regeneration and transformation (root expiant system).- 9. Somatic embryogenesis in carrot.- 10. Low density cultures: microdroplets and single-cell nurse cultures.- 11. Tobacco protoplast isolation, culture and regeneration.- Section B: Tissue Culture & Transformation of Crop Species.- 1. Embryogenic callus, cell suspension and protoplast cultures of cereals.- 2. Transformation and regeneration of rice protoplasts.- 3. Transformation and regeneration of orchardgrass protoplasts.- 4. Transformation and regeneration of oilseed rape protoplasts.- 5. Regeneration and transformation of potato by Agrobacterium tumefaciens.- 6. Transformation of tomato with Agrobacterium tumefaciens.- 7. Regeneration and transformation of sugarbeet by Agrobacterium tumefaciens.- 8. Regeneration and transformation of apple (Malus pumila Mill.).- 9. Transformation and regeneration of maize protoplasts.- 10. Regeneration and transformation of barley protoplasts.- 11. Agrobacterium-mediated transformation of potato stem and tuber tissue, regeneration and PCR screening for transformation.- 12. Production of fertile transgenic wheat by microprojectile bombardment.- 13. Transient gene expression and stable genetic transformation into conifer tissues by microprojectile bombardment.- Section C: Propagation &Conservation of Germplasm.- 1. Clonal propagation of orchids.- 2. Clonal propagation of palms.- 3. Clonal propagation of conifers.- 4. Cytological techniques.- 5. Restriction fragment analysis of somaclones.- 6. Virus elimination and testing.- 7. Clonal propagation of Citrus.- 8. Clonal propagation of eucalypts.- 9. Cryopreservation of plant tissue cultures: the example of embryogénie tissue cultures from conifers.- Section D: Direct Gene Transfer & Protoplast Fusion.- 1. Gene transfer by particle bombardment.- 2. Transformation of pollen by particle bombardment.- 3. Electrical fusion of protoplasts.- 4. Cybrid production and selection.- 5. Fluorescence-activated analysis and sorting of protoplasts and somatic hybrids.- 6. RFLP analysis of organellar genomes in somatic hybrids.- 7. Isolation and uptake of plant nucle.- 8. In situ hybridization to plant metaphase chromosomes: Radioactive and non-radioactive detection of repetitive and low copy number genes.- 9. Chemical fusion of protoplasts.- Section E: Reproductive Tissues.- 1. In vitro fertilisation of maize.- 2. Endosperm culture.- 3. Endosperm culture.- 4. Hybrid embryo rescue.- 5. In vitro culture of Brassica juncea zygotic proembryos.- Section F: Mutant Selection.- 1. Use of chemical and physical mutagens in vitro.- 2. In vitro culture, mutant selection, genetic analysis and transformation of Physcomitrella patens.- Section G: Secondary Metabolites.- 1. Tropane alkaloid biosynthesis in vitro.- 2. Anthocyanin biosynthesis in vitro.- 3. Biosynthesis of monoterpene indole alkaloids in vitro.- Section H: Tissue Culture Techniques for Fundamental Studies.- 1. Establishment of photoautotrophic cell cultures.- 2. Zinnia mesophyll culture system to study xylogenesis.- 3. Cell cycle studies: induction of synchrony insuspension cultures of Catharanthus roseus cells H. Kodama.- 4. Thin Cell Layer (TCL) method to programme morphogenetic patterns.- 5. In vitro infection of Arabidopsis with nematodes.