Washing and Sterilization in Plant Tissue Culture, Bhojwani, Razdan, Calcium Hypochlorite Solution, UV-Sterilization, Plant Material, Transfer of Plant

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Home >> Culture Media and Cell Culture >>Washing and Sterilization in Plant Tissue Culture

Culture Media and Cell Culture
Laboratory Space Requirement for Plant Tissue Culture
Washing and Sterilization in Plant Tissue Culture
Culture Media
Suspension Cultures
Culture of Single Cells (Bergmann's Cell Plating Technique)
Use of Cell Culture in Mutant Selection
Use of Cell Culture in Production of Secondary Metabolites (Suspension Cultures and Immobilized Plant Cells)

	Washing and Sterilization in Plant Tissue Culture - New glassware is always washed using detergents especially designed for the purpose to remove all traces of acids, etc. Finally the glassware is rinsed in tap water and then in distilled water. Sometimes; the glassware may need to be autoclaved before washing to remove agar medium and to destroy microbial contaminants.
The culture medium (details of which are given in the next section), and the glassware are autoclaved at 100°C far 20 min. If autoclave is not available, a pressure cooker may be used Growth factors, such as GA3, zeatin, ABA, urea, etc. (which are thermolabile) should not be autoclaved, but sterilized separately by membrane filtration (far details consult Bhojwani and Razdan, 1983). Instruments, such as forceps, scalpels, needles, and spatula are sterilized by dipping in 95% ethanol fallowed by flaming and cooling. Plant material is surface sterilized using sodium or calcium hypochlorite solution (0.3 - 0.6%) far 15 -30 min. Far ovule, embryo. and end as perm culture, only the seeds are surface sterilized and the ovule or embryo or endosperm is dissected under aseptic conditions (usually in a laminar air flaw cabinet).
Similarly far shoot apices, the buds are surface sterilized and the apices are dissected under aseptic conditions. After surface sterilization, plant material is rinsed 3-4 times in sterile distilled water.All operations including transfer of plant material are carried out under aseptic conditions, preferably under the hood of a laminar air flaw cabinet, which are commercially available far tissue culture laboratories. These cabinets have now replaced UV- sterilization wooden cabinets earlier utilised far this purpose.

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