Marker Genes, Transformed Cells, Calli, Kanamycin, Screenable Genes, Scoreable Genes, Reporter Genes, Antibiotic, Herbicides

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Gene Transfer Methods in Plants Gene Transfer Methods in Plants Introduction Target Cells for Transformation Theoretical Routes for Production of Transgenic Plants Vectors of Gene Transfer Vectors based on Ti and Ri Plasmids of Agrobacterium Structure and Function of Ti and Ri Plasmids Selectable Marker Genes in Vectors Used for Gene Transfer T-DNA Transfer Process Vectors based on Ti and Ri Plasmids Virus Vectors Transformation Techniques Using Agrobacterium Prerequisites for Production of Transgenic Plants Explants Used for Transformation Marker Genes for Selection and Scoring of Tranformed Cells/Calli or Shoots Reporter Genes Used as Scoreable Markers and their Enzyme Assays Neomycin Phosphotransferase Gene (nptII) Chloramphenicol Acetyl Transferase Gene (cat)

Home >> Plant Biotechnology and Genomics >> Gene Transfer Methods in Plants >>Marker Genes for Selection and Scoring of Transformed Cells/Calli or Shoots

Marker Genes for Selection and Scoring of Transformed Cells/Calli or Shoots
After explants are inoculated with Agrobacteriumcarrying the requisite vector having the gene of interest, we need to select the transformed cells/tissues. This is facilitated by the presence of selectable marker genes available in the vector. The selectable marker genes enable the transformed cells to survive in media containing toxic levels of the selection agent, which is usually an antibiotic or a herbicide. Tobacco is used as a model transformation system, where explants (e.g. leaf discs) are placed on regeneration medium containing an antibiotic like kanamycin and transformed shoots can be obtained directly. Any cells, which are not transformed, die due to the presence of kanamycin. Other antibiotics and herbicides may require more judicious use, since even low concentrations can cause rapid cell death. In some cases, selection is exercised only after the regeneration is achieved, because adventitious root formation is sensitive to antibiotics.

Following successful selection, the next requirement is the regeneration of shoots from transformed calli. Since the explants may be heterogeneous and non-transformed tissue may escape initial rejection, several measures are used to ensure that very few non-transformed shoots escape the rejection. The use of leaf discs ensures this, because most cells at the wounded edges, which, undergo regeneration, are in contact with the selection agent.

Screening techniques, also offer an alternative in which no selection pressure is imposed on cells or shoots developing from inoculated explants. In this case, samples of tissues are taken from all regenerated shoots and tested for the expression of a marker gene. A number of marker genes, described as screenable genes or scoreable genes or reporter genes are listed in. The
most commonly used reporter genes are nptll, cat, gus, lux, and pmi, which will be briefly described.

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β-Glucuronidase (GUS)Gene (uidA) Luciferase Gene (lux) Green Fluorescent Protein Gene Phosphomannose Isomerase Gene (pmi) Agrobacterium-Mediated Transformation in Monocots Agroinfection and Gene Transfer Physical Delivery or DNA Mediated Gene Transfer (DMGT) Chemically Stimulated DNA Uptake by Protoplasts Microinjection and Macroinjection Microprojectiles for Gene Transfer Electroporation Method for Gene Transfer Liposome Mediated Gene Transfer Calcium Phosphate Precipitation Method Transformation Using Pollen or Pollen Tube Incubation of Dry Seeds,Embryos,Tissues or Cell in DNA Transformation by Ultrasonication Removal of Selectable Marker Gene from Transgenic Plants Need for Removal of Selectable Marker Gene Genetic Systems Used for Removal of Marker Gene Examples of North American Plant Transformation Facilities Plant Transformation Facilities (PTFs)

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