Team:Warsaw/Calendar-Main/22 August 2008

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Tests for ampicillin resistance given by protein added to medium

Piotr

Purification Omp_alpha of antibiotic resistance contamination:
Transformation of E. coli TOP10 strain with pACTC177+OmpA_alpha.

Cloning of protein A DNA to pET15b+OmpA-alpha plasmid

Antoni

Control digest of pGeneart+A plasmids with NdeI and NotI (Orange buffer).
Gel elctrophoresis. Choice of proper clone.
Digest of pGeneart+A with NdeI and NotI (Orange buffer).
Gel electrophoresis of digested plasmid and gel-out of 470 bp band.
Overnight ligation of pET15b+alpha and A (pET15b+alpha from 19 August).

Cloning of alpha to pACYC177+OmpA_A_omega and pACYC177+OmpA_Z_omega

Michał K.

Ligation of digested vectors from 14 August and alpha DNA fragment from 19 August (more vectors added).
Electroporation of E. coli TOP10 with ligations products.
Transformants plating on LB + kanamycin.

@@ Line 8: / Line 8: @@
 <h3>Cloning of protein A DNA to <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2BOmpA-alpha>pET15b+OmpA-alpha</a> plasmid</h3><h4>Antoni</h4>
 <p><ol>
-<li> <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation">Isolation of plasmids</a> from cultures inocluated on <a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/21_August_2008">previous day</a> (<A href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pGeneart-A>pGeneart+A</a> and <A href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2BOmpA-alpha>pET15b+OmpA-alpha</a>). </li>
 <li> Control <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#digest">digest</a> of <A href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pGeneart-A>pGeneart+A</a> plasmids with NdeI and NotI (Orange buffer). </li>
 <li>Gel elctrophoresis. Choice of proper clone. </li>
-<li> <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#digest">Digest</a> <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2BOmpA-alpha>pET15b+OmpA_alpha</a> and <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pGeneart-A>pGeneart+A</a> with NdeI and NotI (Orange buffer), <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2BOmpA-alpha>pET15b+OmpA_alpha</a><a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#removing"> dephosphorylation with CIAP</a>. </li> </li>
+<li> <a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#digest">Digest</a> of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pGeneart-A>pGeneart+A</a> with NdeI and NotI (Orange buffer). </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 (470 bp for protein A lane and 6200 for pET15b lane). </li>
+<li> Gel electrophoresis of digested plasmid and <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_isolation_from_agarose_gel">gel-out</a> of 470 bp band. </li>
-<li> Overnight <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#ligation">ligation</a> of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2BA-alpha>pET15b+alpha and A</a>. </li>
+<li> Overnight <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#ligation">ligation</a> of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pET15b%2BA-alpha>pET15b+alpha and A</a> (pET15b+alpha from <a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/19_August_2008">19 August</a>). </li>
 </ol></p>
 <h3>Cloning of alpha to <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-A-omega>pACYC177+OmpA_A_omega</a> and <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-Z-omega>pACYC177+OmpA_Z_omega</a></h3><h4>Michał K.</h4>