Team:Warsaw/Calendar-Main/29 July 2008

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1. Separate transformant colonies (transformation from previous day) inoculated to liquid LB with kanamycin.
2. Inoculation of pACYC177+OmpA_alpha and pACYC177+OmpA_omega - liquid LB with kanamycin.

Piotr:

Checking if OmpA_omega_A_alpha gives ampicillin resistance.

Inoculation OmpA_omega_A_alpha from various IPTG concentrations (0, 0.1, 0.25, 0.5, 0.75, 1) into the same IPTG concentrations, but with various ampicillin concentrations(25, 50, 75, 100) in relation: 1:50.

Measurement of bacterial culture growth (OD) in the evening:
ampicillin concentration (μg/mL):IPTG concentration (mmol/mL):
00,10,250,50,751
251,5581,4691,5871,491,5661,311
501,4251,4351,5241,0550,9200,935
751.090,9891.4470.9710.9510.992
1000.090,6851.3781.0780.9770.992

Inoculation of OmpA_omega_A_alpha into various IPTG concentrations: 0, 0.1, 0.25, 0.5, 0.75 (repeat is needed!!!)


Checking OmpA_omega_A_alpha and OmpA_A_alpha expression.
  1. Spinnign
  2. Suspending
  3. Adding of lysis buffer
  4. Boiling
  5. Putting into poliacrylamide gel
  6. Transfer onto nitrocellulose
  7. Blocking
  8. Anti-A antibody binding
  9. Washing
  10. Anti-rabbit antibody binding
  11. Developing with BCIP and NBT
(Zdjęcie żelu)

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