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Escherichia coli recA  −  strains are usually used for cloning to prevent insert instability  via  RecA-dependent recombination. Here, we report that  E. coli  BW25113 ( recA  + ) competent cells prepared by using a previously reported transformation and storage solution (TSS) had 100-fold or higher transformation efficiency than the commonly used  E. coli  cloning strains, including XL1-Blue MRF’. The cloning success rates with  E. coli  BW25113 were 440 to 1,267-fold higher than those with  E. coli  XL1-Blue MRF’ when several inserts were assembled into four vectors by using a simple DNA assembly method. The difference was in part due to RecA, as the  recA  deletion in  E. coli  BW25113 reduced the transformation efficiency by 16 folds and cloning success rate by about 10 folds. However, the transformation efficiency and the cloning success rate of the  recA  deletion mutant of  E. coli  BW25113 are still 12- and &amp;gt;48-fold higher than those of  E. coli  XL1-Blue MRF’, which is a commonly used cloning strain. The cloned inserts with different lengths of homologous sequences were assembled into four vectors and transformed into  E. coli  BW25113, and they were stably maintained in BW25113. Thus, we recommend using  E. coli  BW25113 for efficient cloning and DNA assembly.

Escherichia coli BW25113 Competent Cells Prepared Using a Simple Chemical Method Have Unmatched Transformation and Cloning Efficiencies