Cloning omega-A fusion on pKS (second attempt)

Michał L., Ewa, Marcin

1. Isolation of plasmids from transformants.
2. Digest of plasmids with SacI and NotI (BamHI buffer).
3. Gel electrophoresis of products of digest.
4. 2 selected clones were send to DNA sequencing lab.

Cloning of protein Z DNA to pET15b-OmpA-omega in place of OmpA

Paweł

Ligations transformed into TOP10 and plated on LB + ampicillin.

Cloning of protein A DNA to OmpA constructs

Michał K.

1. Colony PCR on colonies from plates with transformations pACYC177+OmpA_A_omega. Primers used: AL+SacI and AP+NotI
2. Confirmed transformant colonies inoculated to liquid LB with kanamycin.
3. Isolation of plasmids from cultures inocluated on previous day (pACYC177+OmpA_A_alpha).
4. Control digest of isolated plasmids with BamHI and SacI.

Antoni

Setup of overnight culture E. coli TOP10 (LB broth) for preparation of chemocompetent bacteria.