Polyethylene Glycol Peg Treatment, Technique gives Frequency, Reproducible Results, Protoplasts from Unrelated, Soybean Tobacco

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Protoplast Culture Regeneration and Somatic Hybridization Protoplast Culture Regeneration and Somatic Hybridization - Introduction Isolation of Proplasts Mechanical Method Enzymatic Method Direct One Step Method Sequential Two Step Method Purification of Isolated Protoplasts Sedimentation and Washing Flotation Viability and Plating Density of Protoplasts Testing Protoplast Viability Minimum Plating Density MPD and Culturing Individual Protaplasts Protaplast Culture and Regeneration of Plants Isolation of Subprotoplasts Protoplast Fusion and Somatic Hybridization Techniques of Protoplast Fusion

Home >> Plant Biotechnology and Genomics >> Protoplast Culture Regeneration and Somatic Hybridization >> Polyethylene Glycol Peg Treatment

	Polyethylene glycol (PEG) treatment. Since 1974, protoplast fusion has been successfully achieved in several crops, using polyethylene glycol (PEG) as a fusogen. The technique gives high frequency of fusion with reproducible results and involves low cytotoxicity. The technique can be used for fusion of protoplasts from unrelated plant taxa (e.g. soybean – tobacco, soybean – maize, and soybean – barley), from unrelated animal taxa and also between those from animal and plant cells.
The agglutination of protoplasts, during PEG-treatment, can be brought about by the following two different methods: (i) When protoplasts are available in sufficient quantities, 1 ml of culture medium with suspended mixture of both the types of protoplasts is added to 1 ml of 56% solution of PEG and the tube shaken for 5 sec. The protoplasts are allowed to sediment for 10 min, washed with growth medium and examined for successful agglutination and fusion. (ii) If protoplasts are available in microquantities, drop cultures can be used.
Two types of protoplasts are mixed in equal quantities and 4-6 microdrops (100 ml each) are placed in a small Petri plate and allowed to settle for 5-10 min. at room temperature. Two to three microdrops (50 ml each) of PEG are added from the periphery in each Petri plate, which is incubated for 30 min at room temperature (24oC). This leads to agglutination of protoplasts. Sometimes a cover glass is placed in the middle of Petri plate before protoplast suspension is poured. This avoids sticking of protoplasts to the floor of Petri plate and also makes it convenient to handle the protoplasts including their fixation, staining and examination. After PEG treatment, protoplasts are gradually washed and during this process, most of the fusion is achieved. PEG is then replaced by culture medium to allow growth of fused protoplasts.

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Induced Fusion NaNo₃-treatment Treatment with Calcium Ions ca⁺⁺ at High pH Polyethylene Glycol PEG Treatment Electrical Fusion Selection of Fused Protaplasts Chromosome Status of Somatic Hybrids Interspecific Hybrids Produced Through Protoplast Fusion Intergeneric Hybrids Produced Through Protaplast Fusion Use of Protoplast Fusion and Somatic Hybrids Somatic Hybrids for Gene Transfer Asymmetry in Somatic Hybrids for Gene Transfer Somatic Hybrids for Cytoplasmic Male Sterility Cytoplasmic Hybrids or Cybrids Genetic Modification Transformation of Protoplasts

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