Calcium requirement for protoplast transfection mediated by polyethylene glycol of Lactobacillus casei by PL-1 Phage DNA.

The effects of some divalent cations on protoplast transfection mediated by polyethylene glycol of Lactobacillus casei ATCC 27092 by PL-1 phage DNA in 50 mM Tris-maleate buffer (pH 6.0) were investigated. The efficiency of transfection increased about 30 times in the presence of 10 mM Ca2+. Sr2+ increased the transfection rate as well, but Ba2+, Mn2+, and Mg2+ did not. Co2+ and Zn2+ inhibited transfection. The simultaneous use of Ca2+ and Mg2+ increased the transfection efficiency. Impairment of transfection caused by lack of Ca2+ could not be reversed by the addition of Ca2+ later. A decrease in the Ca2+ concentration to an ineffective level before transfection ended immediately inhibited transfection. Protoplasts were transfected with a phage adsorption mutant resistant to PL-1, also, and these metal ions had the same effect. Multiplication of phages in the transfected protoplasts was independent of the presence or absence of calcium ions. Calcium ions seemed to be involved in the entry of PL-1 DNA into the host protoplasts.