Cloning of truncated fragment of protein A

Piotr

Inoculation of some pACYC177+OmpA_alpha + deltaA and pACYC177+OmpA_omega + deltaA colonies of tranformnts.

Checking the expression of omp_omega_A_alpha and omp_A_alpha

Piotr

Inoculation of omp_omega_A_alpha and omp_A_alpha with inductor (0,5 mmol/mL IPTG) and negative control without IPTG.

Checking if omp_omega_A_alpha gives ampicillin resistance

Piotr

Inoculation of omp_omega_A_alpha into various IPTG concentrations: 0, 0.1, 0.25, 0.5, 0.75, 1 mmol/mL.

Preparing pACYC177+OmpA_omega_deltaA construct

Michał K.

1. Digest of pACYC177+OmpA_omega_deltaA_alpha (from 25 July) with BamHI and NotI (BamHI buffer). DNA ends blunting blunting with Klenow fragment (3 hr).
2. Gel elctrophoresis and gel-out of proper band - 4300 bp.
3. Overnight ligation of isolated DNA fragment (1 hr).
4. Transformation of E. coli TOP10 strain with ligation.
5. Transformants plating on LB + kanamycin.