Team:Warsaw/Calendar-Main/18 September 2008
From 2008.igem.org
(Difference between revisions)
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primers (annealing temperature 65 °C; elongation length 60s) to obtain <a href=http://partsregistry.org/wiki/index.php?title=Part:BBa_K103009>linker_alpha (BBa_K103009)</a> fragment. </li> | primers (annealing temperature 65 °C; elongation length 60s) to obtain <a href=http://partsregistry.org/wiki/index.php?title=Part:BBa_K103009>linker_alpha (BBa_K103009)</a> fragment. </li> | ||
- | <li> Gel electrophoresis of PCR product and <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_isolation_from_agarose_gel">gel-out</a> of proper bands (linker_alpha - 600 bp).</li> | + | <li> Gel electrophoresis of PCR product <a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/18_September_2008#fig3">Fig. 3</a> and <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_isolation_from_agarose_gel">gel-out</a> of proper bands (linker_alpha - 600 bp).</li> |
<li><a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#digest">Digest</a> of purified PCR product with EcoRI and BcuI (BamHI buffer). </li> | <li><a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#digest">Digest</a> of purified PCR product with EcoRI and BcuI (BamHI buffer). </li> | ||
<li><a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_purification_after_enzymatic_reaction">Clean-up</a> of digested PCR product.</li> | <li><a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_purification_after_enzymatic_reaction">Clean-up</a> of digested PCR product.</li> | ||
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<li> Gel electrophoresis of PCR products and <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_isolation_from_agarose_gel">gel-out</a> of proper bands (ΔA - 250 bp, linker_alpha - 600 bp, linker_omega - 350 bp and OmpA_omega - 900 bp).</li> | <li> Gel electrophoresis of PCR products and <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_isolation_from_agarose_gel">gel-out</a> of proper bands (ΔA - 250 bp, linker_alpha - 600 bp, linker_omega - 350 bp and OmpA_omega - 900 bp).</li> | ||
- | <li><a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#digest">Digest</a> of purified PCR products and <a href=http://partsregistry.org/wiki/index.php?title=Part:pSB1A3>pSB1A3</a> standard plasmid with EcoRI and BcuI (BamHI buffer) and <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2BOmpA-alpha>pET15b+OmpA_alpha</a> plasmid with NdeI and SacI (BamHI buffer). <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#removing">Dephosphorylation</a> (CIAP) of plasmids.</li> | + | <li><a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#digest">Digest</a> of purified PCR products and <a href=http://partsregistry.org/wiki/index.php?title=Part:pSB1A3>pSB1A3</a> standard plasmid with EcoRI and BcuI (BamHI buffer) and <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2BOmpA-alpha>pET15b+OmpA_alpha</a> plasmid with NdeI and SacI (BamHI buffer) <a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/18_September_2008#fig4">Fig. 4</a>. <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#removing">Dephosphorylation</a> (CIAP) of plasmids.</li> |
<li><a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_purification_after_enzymatic_reaction">Clean-up</a> of digested PCR products.</li> | <li><a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_purification_after_enzymatic_reaction">Clean-up</a> of digested PCR products.</li> | ||
- | <li> Gel electrophoresis of digested plasmids and <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_isolation_from_agarose_gel">gel-out</a> of proper bands (OmpA_link - 500 bp and <a href=http://partsregistry.org/wiki/index.php?title=Part:pSB1A3>pSB1A3</a> - 2200 bp).</li> | + | <li> Gel electrophoresis of digested plasmids <a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/18_September_2008#fig5">Fig. 5</a> and <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_isolation_from_agarose_gel">gel-out</a> of proper bands (OmpA_link - 500 bp and <a href=http://partsregistry.org/wiki/index.php?title=Part:pSB1A3>pSB1A3</a> - 2200 bp).</li> |
<a name="fig1"><img src="https://static.igem.org/mediawiki/2008/c/c3/Grad2_17_09.jpg" width=300/></a><var><b>Fig. 1. Gradient PCR (temperatures:55-75 °C)</b><br> | <a name="fig1"><img src="https://static.igem.org/mediawiki/2008/c/c3/Grad2_17_09.jpg" width=300/></a><var><b>Fig. 1. Gradient PCR (temperatures:55-75 °C)</b><br> | ||
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4. ompA_omega<br></var> | 4. ompA_omega<br></var> | ||
+ | <a name="fig3"><img src="https://static.igem.org/mediawiki/2008/d/d0/Go_17_09.jpg" width=300/></a><var><b>Fig. 3. PCR to obtain linker_alpha</b><br> | ||
+ | 1. Marker<br> | ||
+ | 2. linker_alpha<br></var> | ||
+ | <a name="fig4"><img src="https://static.igem.org/mediawiki/2008/6/6a/Go_29_09.jpg" width=300/></a><var><b>Fig. 4. Control SacI/NdeI digest of pET15b_OmpA_omega </b><br> | ||
+ | 1. Marker<br> | ||
+ | 2. digested pSB1A3<br></var> | ||
+ | |||
+ | <a name="fig5"><img src="https://static.igem.org/mediawiki/2008/9/96/Go2_19_09.jpg" width=300/></a><var><b>Fig. 5. Control EcoRI/BcuI digest of pSB1A3 </b><br> | ||
+ | 1. Marker<br> | ||
+ | 2. digested pSB1A3<br></var> | ||
</html> | </html> |
Revision as of 22:56, 26 October 2008
'Hunter and prey' system tests: Competition testsWeronikaInoculation of pACYC177+OmpA_A_alpha, pACYC177+OmpA_Z_omega, pACYC177+OmpA_A_omega and pACYC177+OmpA_Z_alpha + induction using 0.25mM iPTG MutD5 testingPiotrInoculation of mutD5 into 100 ml of LB and rendering them chemocompetent. Electroporation with pACYC177+OmpA_Z_alpha, pACYC177+OmpA_Z_omega, pACYC177+OmpA_A_omega and pACYC177+OmpA_omega_ΔA. Optimisation of primers for preparation of partsMichał K.
Preparation of pSB1A3 carrying ΔA (BBa_K103003)Michał K.
Preparation of pSB2K3 + _alphaMichał K.
Preparation of pSB2K3 + _omegaMichał K.
Preparation of BioBricksMichał K.
1. Marker 2. 55 °C linker_alpha 3. 60 °C linker_alpha 4. 65 °C linker_alpha 5. 70 °C linker_alpha Fig. 2. PCR to obtain inserts 1. Marker 2. deltaA 3 omega 4. ompA_omega Fig. 3. PCR to obtain linker_alpha 1. Marker 2. linker_alpha Fig. 4. Control SacI/NdeI digest of pET15b_OmpA_omega 1. Marker 2. digested pSB1A3 Fig. 5. Control EcoRI/BcuI digest of pSB1A3 1. Marker 2. digested pSB1A3
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