Team:Warsaw/Calendar-Main/18 June 2008

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Revision as of 17:15, 12 October 2008


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Isolation of pACYC177.
Digest of purified pACYC177 with NdeI and BamHI (Tango 2x buffer).
Gel electrophoresis (Fig. 2) and isolation of proper band (3200 bp).
Clean-up of OmpA_alpha and OmpA_omega digest products (Fig. 1).
Ligation of purified DNA (pACYC177 with OmpA_alpha and OmpA_omega DNA fragments).
Transformation of E.coli TOP10 with ligation products.
Plating transformants on LB+tetracycline.

Counting of blue colonies:

Results of rifampicin test
Strain	Inducer / repressor	Number of colonies on rifampicin
Top10	none	8
Top10 pMPM-AID	0.1% Arabinose	72
Top10 pMPM-AID	none	4
Top10 pMPM-A+T13	0.1% Arabinose	148
Top10 pMPM-A+T13	none	6
Top10 pMPM-AT6	0.1% Arabinose	21
Top10 pMPM-AT6	none	27

Fig. 1. OmpA_alpha (lane 2) and OmpA_omega (lane 3) digest products.

Fig. 2. pACYC177 digested with NdeI and BamHI (lane 2).

@@ Line 7: / Line 7: @@
 <h3>Preparation of constructs with OmpA protein fusions</h3>
 <h4>Michał K.</h4>
-<ol><li><a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation">Isolation</a> of pACYC177.</li>
+<ol><li><a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#plasmid_DNA_isolation">Isolation</a> of <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177>pACYC177</a>.</li>
 <li><a href="https://2008.igem.org/Wiki/Team:Warsaw/igem_notebook.htm#digest">Digest</a> of purified <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177>pACYC177</a> with NdeI and BamHI (Tango 2x buffer). </li>
 <li> Gel electrophoresis (<a href="https://2008.igem.org/wiki/index.php?title=Team:Warsaw/Calendar-Main/18_June_2008#fig2">Fig. 2</a>) and <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#DNA_purification_after_enzymatic_reaction">isolation</a> of proper band (3200 bp).</li>