[DIYbio] Re: Glass bead transformation

It seems pretty convenient. In the book they didn't make protoplasts at first. It seemed to penetrate cell wall anyway.

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3768579/

"Glass bead transformation of Gram positive bacteria was conducted as follows. Aliquots of 0.5 ml of protoplast suspension were placed in 15 ml conical disposable polypropylene centrifuge tubes (Corning, USA). One µg of pGK12 was added to the protoplasts, followed immediately by the addition of 500 µl of 30% polyethylene glycol 6,000 (PEG 6,000) and 0.3 g of acid washed glass beads (212-300 µm in diameter, Sigma, USA). Protoplasts were agitated at the highest speed on a vortex mixer for 15 sec, and then diluted by the addition of 10 ml of transformation buffer. After the beads were allowed to settle, agitated protoplast suspension was transferred to a new 15 ml conical tube. Cells were pelleted by centrifugation at 5,000 xg for 5 min and suspended in 1 ml of BHI supplemented with 0.5 M sorbitol. After incubation at 37°C for 1 h, transformants were recovered by plating the protoplast culture onto BHI agar containing 0.5 M sorbitol and erythromycin at the final concentration of 3 µg/ml and incubated at 37°C for 5 to 7 days"