An improved procedure for the transformation of LiAc treated yeast cells, using single stranded carrier DNA has been previously described (1). Here we describe further improvements and streamlining of this method, yielding reproducibly 106 transformants per microgram of vector DNA, a ten fold improvement, and reducing the time required by 1.5 hours. In addition, the method has been scaled down to various levels, and a colony procedure has been developed. Incubating transformed yeast cells in liquid YPAD for one hour prior to plating on selective medium, increased the transformation efficiency about 7-fold (Table 1). Therefore, we investigated whether any of the incubation steps during transformation had an adverse effect and could be omitted or reduced. We found that it is not necessary to preincubate cells in TE/LiAc and that PEG/TE/LiAc can be added immediately after the cells are aliquoted into tubes containing vector and carrier DNA (see Figure 1). This shortened procedure resulted reproducibly in a transformation frequency of up to 1.2 x 106 colonies per yg of plasmid DNA (Table 1) and this frequency is not increased by post-incubation in YPAD. The volumeshave also been scaled down compared to the original protocol (1). We now routinely regrow to 1 x 107 cells/mlin 50 1l and transform five 50 Al aliquots containing 1 x 108 cells. To transform many strains, 10 ml cultures can be harvested, washed, and resuspended in 50 td ofTE/LiAc; 25 1d samples of thissuspension can be used for control as well as for the transformation, half the volumes of vector, carrier and PEG/TE/LiAc given in Figure 1 are added. This gave the same high transformation efficiency (Table 1). Furthermore, the following colony procedure was developed:'A large size colony, or better several medium size colonies (about 108 cells) are scraped offa YPADplate, washed in 1 ml of sterile water in a microfuge tube, resuspended into 50, l of 1 x LiAc/TE and transformed as in steps 5 to 11 inFigure 1. This procedure which takes only about one hour gives up to 1.5 x 104 transformantsper isg of vector DNA. The high efficiency of the previously developed transformation method (1) made certain applications to some of the most important needs ofcurrent yeast molecularbiology possible (2).