Team:Warsaw/Calendar-Main/3 September 2008

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(Difference between revisions)
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<p><ol><li>Samples were resuspended in PBS, sonicated and centrifuged.</li>  
<p><ol><li>Samples were resuspended in PBS, sonicated and centrifuged.</li>  
<li>Lysis buffer was added separately to pellet and supernatant, then samples were boiled for 10 min. </li>
<li>Lysis buffer was added separately to pellet and supernatant, then samples were boiled for 10 min. </li>
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<li>Lysates loaded on 12% polyacrylamide gel (Fig. 1.) (amount relating to 100 μl of OD=1.0 culture).</li>
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<li>Lysates loaded on 12% polyacrylamide gel (<a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/3_September_2008#fig1">Fig. 1</a>) (amount relating to 100 μl of OD=1.0 culture).</li>
<li>Gel stained with Coomassie Blue. Optimal induction conditions chosen.</li>
<li>Gel stained with Coomassie Blue. Optimal induction conditions chosen.</li>
<li>A-alpha inoculated for overnight culture (Rosetta strain).</li></ol></p>
<li>A-alpha inoculated for overnight culture (Rosetta strain).</li></ol></p>
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<img src="https://static.igem.org/mediawiki/2008/b/bd/September_3_rd.jpg" width=300 /><var>Fig. 1. <b>Supernatants from A-alpha induction with various IPTG concentrations<br>
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<a name="fig1"><img src="https://static.igem.org/mediawiki/2008/b/bd/September_3_rd.jpg" width=300 /></a><var>Fig. 1. <b>Supernatants from A-alpha induction with various IPTG concentrations<br>
The arrow shows place of our overexpressed protein:</b><br>  
The arrow shows place of our overexpressed protein:</b><br>  
1.  22&deg;C 0.5 mM IPTG 5 h<br>
1.  22&deg;C 0.5 mM IPTG 5 h<br>

Revision as of 18:00, 26 October 2008

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Purification of proteins: A-alpha

Piotr, Emilia

  1. Samples were resuspended in PBS, sonicated and centrifuged.
  2. Lysis buffer was added separately to pellet and supernatant, then samples were boiled for 10 min.
  3. Lysates loaded on 12% polyacrylamide gel (Fig. 1) (amount relating to 100 μl of OD=1.0 culture).
  4. Gel stained with Coomassie Blue. Optimal induction conditions chosen.
  5. A-alpha inoculated for overnight culture (Rosetta strain).

Fig. 1. Supernatants from A-alpha induction with various IPTG concentrations
The arrow shows place of our overexpressed protein:
1. 22°C 0.5 mM IPTG 5 h
2. Marker
3. 22°C 0.5 mM IPTG overnight
4. 22°C 1 mM IPTG 5 h
5. 22°C 1 mM IPTG overnight
6. 37°C 0.5 mM IPTG 5 h
7. 37°C 0.5 mM IPTG overnight
8. 37°C 1 mM IPTG 5 h
9. 37°C 1 mM IPTG overnight
10. Marker


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