Team:Warsaw/Calendar-Main/12 July 2008
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+ | <h3>Preparation of constructs with OmpA protein fusions''' and '''Cloning of protein Z DNA to OmpA constructs</h3> | ||
+ | <p><ol> | ||
+ | <li> Isolation of plasmids from cultures inocluated on previous day (pCACYC177 + OmpA_alpha and pACYC177+OmpA_Z_omega). </li> | ||
+ | <li> Control digestation of isolated plasmids with BamHI and SacI (we found good clones for both ligations).</li> | ||
+ | </ol></p> | ||
- | < | + | <h3>Linked PCR Omega-A</h3> |
- | + | <p> | |
- | + | <ul> | |
- | + | <li>reisolated PCR product PCR-omega - 4 µl<br> | |
- | < | + | <li>reisolated PCR product A-homo - 13.5 µl<br> |
- | + | <li>primer | |
- | + | <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#OmegaL+SacI_N">OmegaL+SacI_N</a> - 2 µl</li> | |
- | + | <li>primer | |
- | reisolated PCR product PCR-omega - 4 µl<br> | + | <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#AP+NotI_N">AP+NotI_N</a> - 2 µl</li> |
- | reisolated PCR product A-homo - 13.5 µl<br> | + | <li>Pfu buffer with Mg2+ - 5 µl</li> |
- | + | <li>dNTPs - 1 µl</li> | |
- | <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#OmegaL+SacI_N">OmegaL+SacI_N</a | + | <li>H2o - 22 µl</li> |
- | + | <li>Program:</li> | |
- | <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#AP+NotI_N">AP+NotI_N</a | + | <ol> |
- | Pfu buffer with Mg2+ - 5 µl< | + | <li> 95°C - 3'</li> |
- | dNTPs - 1 µl< | + | <li> 95°C - 30"</li> |
- | H2o - 22 µl< | + | <li> 55°C - 45"</li> |
- | < | + | <li> 68°C - 1'</li> |
- | + | <li> go to step 2 25 x</li> | |
- | < | + | <li> 68° - 10'<br> |
- | + | <li> keep in 4°</li> | |
- | + | </ol></li> | |
- | + | <li>gel electrophoresis</li></ul> | |
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- | gel electrophoresis< | + | |
</p> | </p> | ||
- | + | </html> | |
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Revision as of 17:53, 1 October 2008
Preparation of constructs with OmpA protein fusions''' and '''Cloning of protein Z DNA to OmpA constructs
Linked PCR Omega-A
|