Escherichia coli XL10-Gold Bacteria Produce Bacteriophage

LETTER

Site-directed mutagenesis (SDM) is a powerful method to introduce specific mutations into DNA sequences (1). Package inserts in commercial kits for SDM sold by Agilent Technologies (QuikChange methods) recommend the use of XL10-Gold ultracompetent bacteria (2). We find, however, that XL10-Gold bacteria produce bacteriophage that resemble ϕ80 phage. The phage have not been fully characterized, but, like ϕ80 phage, they replicate lytically or they lysogenize host bacteria (Fig. 1). In addition, infection requires the bacterial fhuA gene, which is the receptor for ϕ80 as well as for T1 and T5 phage (3). Phage titers of ∼10⁸ phage/ml were observed from cultures of XL10-Gold bacteria from a 7-year-old stock stored at −80°C as well as from recently purchased competent cells. We also observed that bacterial cells in liquid culture aggregated as expected from phage production and cell lysis.

Fig 1.

Plaques formed by bacteriophage produced in cultures of XL10-Gold cells. XL10-Gold bacteria from purchased competent cells were cultured in Luria broth (LB). A few drops of chloroform were added to the bacterial culture, the culture was subjected to a gentle vortexing procedure, and the debris was pelleted by centrifugation. The supernatant was diluted, and the dilutions were mixed with 2.5 ml of soft agar containing 0.05 ml permissive plating bacteria; each mixture was poured over the surface of an LB plate to form a top agar layer. Plates were incubated at 30°C for 18 h.

Although bacteriophage may not be a problem for many applications, XL10-Gold bacteria should be avoided in microbiology laboratories. It is noteworthy that bacteriophage-like DNA sequences have been detected in commercial preparations of the Taq DNA polymerase (4). Bacteriophage production compromises bacterial growth, and phage DNA has the potential to produce misleading results.