
%0 Journal Article
%A Sharma, Anubhuti 
%A Girdhar, Arushi 
%A Srivastava, Nidhi 
%T Development of strategy for competent cell preparation and high efficiency plasmid transformation using different methods
%J The South Pacific Journal of Natural and Applied Sciences
%D 2012/02/13
%V 29
%N 1
%P 17-20
%K Competent cells, ligation, molecular cloning, transformation.
%R https://doi.org/10.1071/SP11004
%U https://www.publish.csiro.au/paper/SP11004 
%X Current cloning technologies based on site-specific recombination are efficient, simple to use, and flexible. With the recent availability of complete genomic sequences of many organisms and plants, high-throughput and cost-efficient systems for gene cloning and functional analysis are in great demand. This study compares two different methods of preparation of competent cells using two strains of E. coli DH5&#945; and HB101. From results the most efficient strain was found DH5&#945; for cloning as it supports blue white screening utilizing galactosidase activity. The concentration of calcium chloride is another important factor; various concentrations of CaCl2 were tried. Optimum concentration was found to be 75 mM. However PEG also has great influence on transformation efficiency, use of 40% PEG gave the best transformation efficiency. A convenient and rapid method for the genetic transformation of E. coli with appropriate plasmid is proposed which can be utilised for high efficiency transformation in normal laboratory conditions. 
