Molecular Cloning Fourth Edition, A Laboratory Manual, by Michael R. Green and Joseph Sambrook

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The Hanahan Method for Preparation and Transformation of Competent E. coli: High-Efficiency Transformation

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Starting in the 1970s and continuing to this day, many variations on the basic technique of bacterial transformation have been described in the literature, directed chiefly toward optimizing the efficiency of transformation of different E. coli strains by plasmids. The variations include using complex cocktails of divalent cations in different buffers, treating cells with reducing agents, adjusting the ingredients of the cocktail to the genetic constitution of particular strains of E. coli, harvesting cells at specific stages of the growth cycle, altering the temperature of growth of the culture before exposure to chemicals, optimizing the extent and temperature of heat shock, freezing and thawing cells, and exposing cells to organic solvents after washing in divalent cations. Using these treatments and others, it is now possible on a routine basis to achieve transformation efficiencies ranging up to 109 transformants/g of superhelical plasmid DNA (for reviews, see Hanahan 1983; Hanahan et al. 1991, 1995; Hanahan and Bloom 1996; Hengen 1996).


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(Limited time special offer.) Molecular Cloning Fourth Edition, A Laboratory Manual, by Michael R. Green and Joseph Sambrook
Molecular Cloning Fourth Edition, A Laboratory Manual, by Michael R. Green and Joseph Sambrook

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