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Team:Warsaw/Calendar-Main/6 August 2008
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<h3>Checking if <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-omega-A-alpha>OmpA_omega_ΔA_alpha</a> gives ampicillin resistance </h3> | <h3>Checking if <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-omega-A-alpha>OmpA_omega_ΔA_alpha</a> gives ampicillin resistance </h3> | ||
| - | <h4>Piotr</h4> | + | <h4>Piotr, Weronika</h4> |
| - | <p>Inoculation of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-omega-A-alpha>OmpA_omega_ΔA_alpha</a> from various IPTG concentrations: 0, 0.1, 0.25, 0.5, 0.75, 1 mM into same IPTG concentrations, but with various ampicillin concentrations (25, 50, 75, 100 mM) in ratio 1:50.</p> | + | <p>Repetition of the test from previous day. Inoculation of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-omega-A-alpha>OmpA_omega_ΔA_alpha</a> from various IPTG concentrations: 0, 0.1, 0.25, 0.5, 0.75, 1 mM into same IPTG concentrations, but with various ampicillin concentrations (25, 50, 75, 100 mM) in ratio 1:50.</p> |
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Checking if OmpA_omega_ΔA_alpha gives ampicillin resistancePiotr, WeronikaRepetition of the test from previous day. Inoculation of OmpA_omega_ΔA_alpha from various IPTG concentrations: 0, 0.1, 0.25, 0.5, 0.75, 1 mM into same IPTG concentrations, but with various ampicillin concentrations (25, 50, 75, 100 mM) in ratio 1:50.
Conclusion: we obtained the best induction using 0.5 - 0.25 mM IPTG. Best ampicillin concentration is 50 - 75 μg/ml. Checking whether degradation of the fusions with OmpA is caused by Lon and OmpT proteases (present in TOP10)EmiliaTest was conducted in E.coli Rosetta strain expressing ompA_omega_ΔA_alpha (with and without induction) and ompA_A_alpha. Transformation of Rosettas with OmpA_omega_ΔA_alpha and OmpA_A_alpha. Preparing pACYC177+OmpA_omega_ΔA constructMichał K.Inoculation of cultures from colonies grown on plate with pACYC177+OmpA_omega_ΔA transformation.
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