Difference between revisions of "Yeast Transformation"
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What’s Transformation? | '''What’s Transformation?''' | ||
In molecular biology, transformation is the genetic alteration of a cell resulting from the direct uptake and incorporation of exogenous DNA (originating outside the cell) from its surroundings through the cell membrane. In yeast, there are numerous ways to perform transformation. One method that is used is Electroporation. | In molecular biology, transformation is the genetic alteration of a cell resulting from the direct uptake and incorporation of exogenous DNA (originating outside the cell) from its surroundings through the cell membrane. In yeast, there are numerous ways to perform transformation. One method that is used is Electroporation. | ||
What's Electroporation? | What's Electroporation?''' | ||
''' | |||
It’s a technique in which an electrical field is applied to cells in order to increase the permeability of the cell membrane, allowing DNA to be introduced into the cell. | |||
'''Important distinctions:''' | |||
Electroporation efficiency = number of transformants per microgram of DNA. | |||
Electroporation frequency= transformation efficiency per viable cells. | |||
Electroporation efficiency = number of transformants per microgram of DNA | |||
Electroporation frequency= transformation efficiency per viable cells | |||
“Freezing of intact cells in sorbitol with calcium at -80C results in higher transformation efficiency by electroporation, giving more than 106 transformants/μg of plasmid DNA after thawing.” | “Freezing of intact cells in sorbitol with calcium at -80C results in higher transformation efficiency by electroporation, giving more than 106 transformants/μg of plasmid DNA after thawing.” | ||
Revision as of 19:29, 22 June 2016
What’s Transformation?
In molecular biology, transformation is the genetic alteration of a cell resulting from the direct uptake and incorporation of exogenous DNA (originating outside the cell) from its surroundings through the cell membrane. In yeast, there are numerous ways to perform transformation. One method that is used is Electroporation.
What's Electroporation? It’s a technique in which an electrical field is applied to cells in order to increase the permeability of the cell membrane, allowing DNA to be introduced into the cell.
Important distinctions:
Electroporation efficiency = number of transformants per microgram of DNA. Electroporation frequency= transformation efficiency per viable cells.
“Freezing of intact cells in sorbitol with calcium at -80C results in higher transformation efficiency by electroporation, giving more than 106 transformants/μg of plasmid DNA after thawing.” Reference: High-efficiency electroporation by freezing intact yeast cells with addition of calcium, by Suga and Hatakeyama.
“Pretreatment of yeast cells with lithium acetate (LiAc) and dithiothreitol (DTT) enhances the frequency of transformation by electroporation. The method shows improvements of 6–67-fold in wild-type strains derived from commonly used Saccharomyces cerevisiae genetic backgrounds. In addition, 15–300-fold improvement in transformation frequency was achieved with several mutant strains of S. cerevisiae that transformed poorly by conventional procedures.”
Reference: An improved protocol for the preparation of yeast cells for transformation by electroporation, by Thompson et al.
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