Team:Warsaw/Calendar-Main/17 September 2008

From 2008.igem.org

(Difference between revisions)
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<h4>Michał K.</h4>
<h4>Michał K.</h4>
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<ol><li><a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#pcr">PCR</a> on pACYC177+OmpA_omega_deltaA_alpha plasmid using  
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<ol><li><a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#pcr">PCR</a> on <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-omega-A-alpha>pACYC177+OmpA_omega_&Delta;A_alpha</a> plasmid using  
<a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#AL_BNXNE">AL_BNXNE</a> and <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#APSacSpe">APSacSpe</a>
<a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#AL_BNXNE">AL_BNXNE</a> and <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#APSacSpe">APSacSpe</a>
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  primers (elongation length 45s) to obtain deltaA fragment. </li>
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  primers (elongation length 45s) to obtain &Delta;A fragment. </li>
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<li> <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#pcr">PCR</a> on pACYC177+OmpA_omega_deltaA_alpha plasmid using  
+
<li> <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#pcr">PCR</a> on <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-omega-A-alpha>pACYC177+OmpA_omega_&Delta;A_alpha</a> plasmid using  
<a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#LinLSXNE">LinLSXNE</a> and <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#AlfaPSpe">AlfaPSpe</a>
<a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#LinLSXNE">LinLSXNE</a> and <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#AlfaPSpe">AlfaPSpe</a>
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primers (elongation length 60s) to obtain link_omega fragment. </li>
primers (elongation length 60s) to obtain link_omega fragment. </li>
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<li><a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#pcr">PCR</a> on pACYC177+OmpA_omega_deltaA_alpha plasmid using
+
<li><a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#pcr">PCR</a> on <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-omega-A-alpha>pACYC177+OmpA_omega_&Delta;A_alpha</a> plasmid using
<a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#OmpLNXNE">OmpLNXNE</a> and <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#LinP_BS">LinP_BS</a>
<a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#OmpLNXNE">OmpLNXNE</a> and <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#LinP_BS">LinP_BS</a>
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primers (elongation length 90s) to obtain Omp_omega fragment. </li>
+
primers (elongation length 90s) to obtain OmpA_omega fragment. </li>
<p>Each reaction was carried out with 25 cycles and in temperature gradient from 55 to 75 &deg;C (four reactions).</p>
<p>Each reaction was carried out with 25 cycles and in temperature gradient from 55 to 75 &deg;C (four reactions).</p>
<li> Gel electrophoresis of PCR products.</li>
<li> Gel electrophoresis of PCR products.</li>

Revision as of 22:45, 25 October 2008

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'Hunter and prey' system tests: Competition tests

Piotr

  1. Plasmid isolation from previous day's cultures.
  2. Digest with SacI and BamHI.
  3. Electrophoresis.

[photo of the gel is to be placed here]

Conclusion: cells with interracting protein survive competition!

MutD5 testing

Emilia

Inoculation of MutD5 into medium with tetracycline.

Optimisation of primers for preparation of parts

Michał K.

  1. PCR on pACYC177+OmpA_omega_ΔA_alpha plasmid using AL_BNXNE and APSacSpe primers (elongation length 45s) to obtain ΔA fragment.
  2. PCR on pACYC177+OmpA_omega_ΔA_alpha plasmid using LinLSXNE and AlfaPSpe primers (elongation length 60s) to obtain link_alpha fragment.
  3. PCR on pACYC177 + OmpA_omega plasmid using LinLSXNE and OmegPSpe primers (elongation length 60s) to obtain link_omega fragment.
  4. PCR on pACYC177+OmpA_omega_ΔA_alpha plasmid using OmpLNXNE and LinP_BS primers (elongation length 90s) to obtain OmpA_omega fragment.

    5. Each reaction was carried out with 25 cycles and in temperature gradient from 55 to 75 °C (four reactions).

  5. Gel electrophoresis of PCR products.

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