Team:Warsaw/Calendar-Main/4 August 2008
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<li>Inoculation of pACYC177+OmpA_alpha and pACYC177+OmpA_omega - liquid LB with kanamycin.</li></ol> | <li>Inoculation of pACYC177+OmpA_alpha and pACYC177+OmpA_omega - liquid LB with kanamycin.</li></ol> | ||
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<h3>Checking for presence of A on the cell membrane</h3> | <h3>Checking for presence of A on the cell membrane</h3> |
Revision as of 22:08, 11 October 2008
Checking the expression of omp_omega_A_alpha and omp_A_alphaPiotrInoculation 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
|
ampicillin concentration (μg/mL): | IPTG concentration (mmol/mL): | |||||
---|---|---|---|---|---|---|
0 | 0.1 | 0.25 | 0.5 | 0.75 | 1 | |
25 | 1.558 | 1.469 | 1.587 | 1.49 | 1.566 | 1.311 |
50 | 1.425 | 1.435 | 1.524 | 1.055 | 0.920 | 0.935 |
75 | 1.09 | 0.989 | 1.447 | 0.971 | 0.951 | 0.992 |
100 | 0.09 | 0.685 | 1.378 | 1.078 | 0.977 | 0.992 |
Inoculation of OmpA_omega_A_alpha into various IPTG concentrations: 0, 0.1, 0.25, 0.5, 0.75 mmol/mL (replication is necessary)
Checking OmpA_omega_A_alpha and OmpA_A_alpha expression
Piotr
- Spinning
- Suspending
- Adding of lysis buffer
- Boiling
- Putting into poliacrylamide gel
- Transfer onto nitrocellulose
- Blocking
- Anti-A antibody binding
- Washing
- Anti-rabbit antibody binding
- Developing with BCIP and NBT
Michał L., Ewa, Marcin
- Separate transformant colonies (transformation from previous day) inoculated to liquid LB with kanamycin.
- Inoculation of pACYC177+OmpA_alpha and pACYC177+OmpA_omega - liquid LB with kanamycin.
Checking for presence of A on the cell membrane
Piotr
Inoculation of omp_A_alfa, omp_Z_alfa and omp_omega_A_alfa (with and without induction).