Plant tissue culture


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Plant tissue culture

  1. 1. Plant tissue culture • Broadly refers to technique of growing plant cells, tissues, organs, seeds or other plant parts in a sterile environment on a nutrient medium Plant tissue culture • The first commercial use of plant propagation on artificial a.k.a. micropropagation media was in the germination and growth of orchid plants in the 1920’s • It was only after the development of a reliable artificial medium by Murashige & Skoog in 1962 that plant tissue culture really took off commercially Why do plant tissue culture? History• Fast commercial propagation of new cultivars • First attempted by Haberlandt (1902) - grew palisade cells – Cost of culture (orchid price decrease) from leaves of various plants but they did not divide• Agriculture • 1934 - White generated continuously growing culture of – Fast selection for crop improvement – nutritional value, meristematic cells of tomato on medium containing salts, pest control, hardiness yeast extract and sucrose and 3 vit B (pyridoxine, thiamine, nicotinic acid) – established the importance of – Cultivation virus free plants additives• Pharmaceuticals – ginseng and taxol • 1953 Miller and Skoog, University of Wisconsin – Madison• Cloning of rare and endangered plants discovered kinetin, a cytokine that plays an active role in• Plant cultures in approved media are easier and safer to organogenesis export Further progress Basis for the cell culture• 1958-60 Morel cultured orchids and dahlias freed them • Plant cells are totipotent from a viral disease • Have the ability to develop into whole plants or plant• 1962 Murashige and Skoog published recipe for M&S organs in vitro when given the correct conditions medium • Not all plant cells are totipotent. However, there are a• 60’s and 70’s Murashige cloned plants in vitro sufficient number of totipotent cells in the plant (e.g. in – raised haploid plants from pollen grains the pith) – used protoplast fusion to hybridize 2 species of tobacco into one plant contained 4N • Differentiated cells have to be dedifferentiated into callus• 70’s and 80’S beginning of genetic engineering and redifferentiated back to somatic embryo that will regenerate the entire plant 1
  2. 2. Culturing (micropropagating) plant Callus tissue - the steps• Is a natural response of the plant tissue to wounding • Selection of the plant tissue (explant) from a healthy• A mass of actively dividing undifferentiated cells produced vigorous ‘mother plant’ - this is often the apical bud, but by plant tissue explant can be other tissue• Cells are totipotent • Establishment of the explant in a culture medium - the• Callus can be medium sustains plant cells growth and encourages cell division – Resuspended in liquid media to create a susupension culture of single totipotent cells – Each plant species (and sometimes the variety within a species) has particular medium requirements that must – Or differentiated into plant with the appropriate be established by trial and error manipulations of culture conditions • Multiplication - the explant gives rise to a callus • Differentiation and organogenesis What is needed? Plant tissue culture media• Explant (some tissues culture better than others) • Mineralals (17 essential elements)• A suitable growth medium • Energy and carbon source - sucrose is preferred• Aseptic conditions, as microorganisms grow much more • Growth regulators – auxins and cytokinins quickly than plant and animal tissue (phytohormones)• Growth regulators - in plants, both auxins & cytokinins • Vitamins• Frequent subculturing to ensure adequate nutrition and to • Organic compounds avoid the build up of waste metabolites • Water • Usually 5.0-5.7 Vitamins Growth regulators• Thiamine Auxins and cytokinins• Pyridoxin • Auxins stimulate cell division in explants• Nicotinic acid – Used for callus induction and growth• Biotin • High concentration suppresses the organized growth and• Citric acid promotes formation of meristem-like cells• Ascorbic acid • IAA, indole-3-acetic acid a light sensitive, natural auxin• Inositol • Cytokinins • Promote cell division and regeneration possibly through the effects on protein synthesis • Zeatin – natural cytokine that induces morphogenesis 2
  3. 3. Homework• In your notebook write a detailed content of a selected plant culture medium. It has to have concentrations of all ingredients!• Make sure you understand what is in it and why. Again, there will be a question about it on the first exam.• Find a name of the plant that is grown on that medium 3