Team:Warsaw/Calendar-Main/29 July 2008

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Revision as of 14:14, 9 October 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):
ampicillin concentration (μg/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 (repeat is needed!!!)

Checking OmpA_omega_A_alpha and OmpA_A_alpha expression.

Spinnign
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

(Zdjęcie żelu)

@@ Line 19: / Line 19: @@
 <table id="result" align="center">
-<tr><th rowspan="2">ampicillin concentration (mikrogramy na mililitr):</th><th colspan="6">IPTG concentration (milimole na mililitr):</td></tr>
+<tr><th rowspan="2">ampicillin concentration (μg/mL):</th><th colspan="6">IPTG concentration (mmol/mL):</td></tr>
 <tr><th>0</th><th>0,1</th><th>0,25</th><th>0,5</th><th>0,75</th><th>1</th></tr>
 <tr><th>25</th><td class="live">1,558</td><td class="live">1,469</td><td class="live">1,587</td><td class="live">1,49</td><td class="live">1,566</td><td class="live">1,311</td></tr>