A Practical Guide for Introducing Gibson Assembly® Gene Constructs into Chemically Competent Cells

The Gibson Assembly ® method is an easy-to-use, robust, seamless cloning method that allows for the efficient cloning of multiple DNA fragments simultaneously. Gene constructs assembled with Gibson Assembly ® are often introduced into E. coli for propagation and maintenance. In this practical guide, we tested three commercially available chemically competent cells with Gibson Assembly ® reaction mixtures. Similar to traditional ligation-dependent cloning methods, salt and enzymes present in the Gibson Assembly ® reaction mixture can be detrimental to competent cells. Although transformation was successful in some cases with percentages as high as 10% of the Gibson Assembly ® reaction mixture, highest transformation efficiencies were generally achieved using lower volumes of the reaction mixture. Based on our results, we recommend using up to 2% of the Gibson Assembly ® reaction as a starting point when transforming chemically competent cells with Gibson Assembly ® constructs.