Bioreactors in Cell Culture

Plant Tissue Culture Application

• Plant tissue culture is used to apply various techniques for plant propagation, genetic improvement, and conservation.
• Different applications, including suspension culture, somatic embryogenesis, organogenesis, haploid production, short-term and long-term germplasm conservation, and somaclonal variations are discussed.

Organogenesis

• Organogenesis, or morphogenesis, is the production of new organs not present in the initial explant.
• Tissues or organs with the capacity for this are called morphogenic or organogenic.
• Adventitious shoots/roots are new organs induced to form on cultured plant tissues.

Somatic Embryogenesis

• Somatic embryogenesis is the process where somatic cells differentiate into embryos.
• Embryos formed by somatic cells are called somatic embryos.
• Somatic embryos can arise directly from explants or indirectly after callusing.
• Somatic embryos resemble zygotic embryos morphologically, are bipolar, and contain typical embryonic organs.
• They are not connected to explant or callus cells by vascular tissue.
• Somatic embryogenesis supports large-scale vegetative propagation and long-term storage of material via cryopreservation.

Callus Culture

• Callus is a mass of unspecialized, unorganized cells produced when explants are cultured on a solid medium supplemented with auxin and cytokinin.
• Callus is useful in producing variations among plantlets.
• Callus cultures may be compact or friable (compact shows densely aggregated cells, friable loosely associated).
• Embryogenic callus leads to plantlet regeneration via organogenesis or embryogenesis.
• Different types of callus exist (embryogenic and non-embryogenic).
• Callus culture involves three stages: induction (cell division), proliferative (rapid cell division), and morphogenesis (differentiation into organized structures).

Cell Suspension Culture

• Friable callus isolated from explants is grown in liquid media for cell suspension culture.
• Agitation (30-100 rpm) is necessary for suspension cultures.
• Suspension cultures grow faster than callus cultures.
• The medium often contains auxin, and pectinase can be added to improve single cell production and growth rate.
• The process of initiation and maintenance of plant cell suspension culture is explained by a diagram.

Haploid Production (Methods)

• Haploid cells have a single set of chromosomes, in contrast to diploid cells (2n).
• Haploid plants are vital for creating homozygous lines and for using plants in breeding programs.
• Methods of haploid production include androgenesis, gynogenesis, distant hybridization, and irradiation effects and chemical treatments.
• Androgenesis uses anther or pollen culture, while gynogenesis uses unfertilized ovules or ovaries.
• In vitro methods for haploid production include androgenesis and gynogenesis.

Comparisons of Anther and Pollen Cultures

• Anther culture is easy, quick and practicable but can have unwanted diploid/aneuploid variation within the plant population from anther sources.
• Pollen culture can overcome these problems.

Gynogenesis

• Gynogenesis is the culture of unfertilized female gametophytes (ovules or ovaries).
• Used when plants don't respond to androgenic methods or when producing albino plants is a problem.
• Gynogenesis is less efficient than androgenesis in some plant families (e.g. Liliaceae, Compositae).

Germplasm Conservation

• Germplasm conservation seeks to preserve genetic diversity of a specific plant or genetic stock.
• Conservation methods include in situ (in the natural habitat) and ex situ (gene banks, seed banks).
• In vitro germplasm conservation methods explained include cryopreservation, cold storage, and low-pressure/low-oxygen storage.
• Protocols for cryopreservation of shoot tips are outlined in a diagram.

Somaclonal Variation

• Somaclonal variation is genetic diversity among plants regenerated from somatic cells (in vitro).
• Although these plants should be clones, some observations show they’re not.
• Somaclonal variation is useful in various breeding programs for useful traits and breeding lines.
• Somaclonal variation can be useful in developing new crop varieties with novel variations.
• Advantages and disadvantages of somaclonal variation are highlighted.