PICHIAPINK-4 Yeast Competent Cells

YCC-018

PICHIAPINK-4 Yeast Competent Cells

100 μl*50

1). Take 5 µg of the plasmid to be transformed, digest and recover.
2). Take a sterile 1.5 ml EP tube and sequentially add 1-5 µg of pre-cooled linear plasmid (volume not higher than 15 µl), 10 µl of yeast transformer, 100 µl of sensory cells melted on ice, 500 µl of Komagataella pastoris transformation solution.
Mix gently by turning 6-8 times.
3). 30°C water bath for 30 min, gently turn the mixture 6-8 times every 10 min.
4). Add 20 μl of dimethyl sulfoxide to each.
5). 42 ℃ water bath for 15 min, every 5 min gently turn and mix 6-8 times.
6). 12,000 rpm instantaneous centrifugation, discard the supernatant add 1 ml of YPD Plus to each cartridge, and resuscitate for 1 h at 30℃.
7). 12,000 rpm instantaneous centrifugation and discard the supernatant, resuspend with 100 μl 0.9% NaCl, coat the plate, and incubate at 30℃ for 48-96 h. The plate was then incubated at 30℃ for 48-96 h. The supernatant was discarded by centrifugation for 6-8 times.

1). Before using the yeast transformer, please boil the tube containing the yeast transformer in boiling water for 5 min, then immediately put it on ice, and store it at -20℃ after use.
2). The transformer solution will precipitate at low temperatures, so please dissolve it completely at room temperature and use it.
3). The whole process of transformation should be aseptic.

100 μl

5 ml

Competent cells are transported on dry ice and stored at -80°C. Yeast transformer needs to be stored at -20°C. Komagataella pastoris transformation solution needs to be stored at 4°C.

6 months