Transformation of intact yeast cells treated with alkali cations

TLDR

The authors investigated transformation of Saccharomyces cerevisiae D13‑1A with plasmid YRp7 using CsCl, showing that a heat pulse combined with polyamines or basic proteins stimulated plasmid DNA uptake. Intact yeast cells treated with alkali cations (Li⁺, Cs⁺, Rb⁺, K⁺, Na⁺) became competent, taking up both circular and linear plasmid DNA (with lower efficiency for linear forms); optimal conditions were 1 h incubation, 5 × 10⁷ cells mL⁻¹, 1.0 M Cs⁺, and polyethylene glycol, yielding transformation efficiencies comparable to conventional protoplast methods for ars1‑containing plasmids but not for 2‑micron origin plasmids.

Abstract

Intact yeast cells treated with alkali cations took up plasmid DNA. Li+, Cs+, Rb+, K+, and Na+ were effective in inducing competence. Conditions for the transformation of Saccharomyces cerevisiae D13-1A with plasmid YRp7 were studied in detail with CsCl. The optimum incubation time was 1 h, and the optimum cell concentration was 5 x 10(7) cells per ml. The optimum concentration of Cs+ was 1.0 M. Transformation efficiency increased with increasing concentrations of plasmid DNA. Polyethylene glycol was absolutely required. Heat pulse and various polyamines or basic proteins stimulated the uptake of plasmid DNA. Besides circular DNA, linear plasmid DNA was also taken up by Cs+-treated yeast cells, although the uptake efficiency was considerably reduced. The transformation efficiency with Cs+ or Li+ was comparable with that of conventional protoplast methods for a plasmid containing ars1, although not for plasmids containing a 2 microns origin replication.