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IJBB Vol.46(5) [October 2009] >


Title: Analysis of comparative efficiencies of different transformation methods of E. coli using two common plasmid vectors
Authors: Roychoudhury, Aryadeep
Basu, Supratim
Sengupta, Dibyendu N
Keywords: Calcium chloride
E. coli
Ethanol
Heat shock
Microwave
Plasmid DNA
Transformation efficiency
Issue Date: Oct-2009
Publisher: CSIR
Abstract: The efficiencies of different transformation methods of E. coli DH5α strain, induced by several cations like Mg2+, Mn2+, Rb+ and especially Ca2+, with or without polyethylene glycol (PEG) and dimethyl sulfoxide (DMSO) were compared using the two commonly used plasmid vectors pCAMBIA1201 and pBI121. The widely used calcium chloride (CaCl2) method appeared to be the most efficient procedure, while rubidium chloride (RbCl) method was the least effective. The improvements in the classical CaCl2 method were found to further augment the transformation efficiency (TR)E for both the vectors like repeated alternate cycles of heat shock, followed by immediate cold, at least up to the third cycle; replacement of the heat shock step by a single microwave pulse and even more by double microwave treatment and administration of combined heat shock-microwave treatments. The pre-treatment of CaCl2-competent cells with 5% (v/v) ethanol, accompanied by single heat shock also triggered the (TR)E, which was further enhanced, when combined heat shock-microwave was applied. The minor alterations or improved approaches in CaCl2 method suggested in the present study may thus find use in more efficient E. coli transformation.
Page(s): 395-400
ISSN: 0975-0959 (Online); 0301-1208 (Print)
Source:IJBB Vol.46(5) [October 2009]

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