Y190 Yeast Competent Cells

YCC-019

Y190 Yeast Competent Cells

100 μl*50

Y190 is a MATa-type yeast strain for GAL4 yeast two-hybrid assay. It can be directly transformed into plasmids or mating with MATa-type yeast strain Y187 for screening. Plasmid pGB was transformed from pGBKT7 with the screening marker TRP1 for expression of DNA-BD fusion proteins with target proteins. Plasmid pACT2 is structurally and functionally similar to pGADT7, and the screening marker is LEU, which is used to express the fusion protein of AD and the target protein.

MATa, ura3- -52, his3- -200, ade2-101, lys2-801, trp1-901, leu2-3, 112, gal4△, gal80△, cyhr2, LYS2:GAL1UAS-HIS3TATA-HIS3, MEL1 URA3::GAL1UAS-GAL1TATA-lacZ

trpl, leu2, cyh2

lacZ, HIS3, MEL1

pGB and pACT2, or pGBKT7 and pGADT7

Transformation efficiency >10⁴ cfu/ug DNA as assayed by pGADT7 plasmid.

1). Take a sterile 1.5 ml EP tube, sequentially add 1-3 μg of pre-cooled target plasmid, 10 μl of Carrier DNA (95-100 ℃ for 5 min in a rapid ice bath, repeat once), and 100 μl of competent cells melted on ice, 500 μl of PEG/LiAc, and gently turn and mix 6-8 times.
2). 30 ℃ water bath for 30 min, every 10 min gently turn over and mix 6-8 times.
3). Add 20 μl of dimethyl sulfoxide to each branch (used to improve the conversion efficiency, can not be done).
4). 42 ℃ water bath for 15 min, every 5 min gently turn over and mix 6-8 times.
5).12,000 rpm instantaneous centrifugation and discard the supernatant, add 1 ml of YPD Plus to each cartridge, and resuscitate at 30℃ for 1 h (used to improve the transformation efficiency, can not be done).
6).12,000 rpm instantaneous centrifugation and discard the supernatant, resuspend with 100 μl of 0.9% NaCl, coat the plate, and incubate at 30℃ for 48-96 hours.

Competent cells are best melted on ice, PEG solution will precipitate at low temperatures, please dissolve completely at room temperature before use.

5 μg/μl, 100 μl

5 ml

Competent cells are transported on dry ice and stored at -80°C. Carrier DNA needs to be stored at -20°C. PEG/LiAc needs to be stored at 4°C.

6 months