Team:Warsaw/Calendar-Main/13 August 2008

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<li>Washing.</li>
<li>Washing.</li>
<li>Anti-rabbit antibody binding.</li>
<li>Anti-rabbit antibody binding.</li>
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<li>Developing with BCIP and NBT (see fig.1.).</li>
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<li>Developing with BCIP and NBT (see <a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/13_August_2008#fig1">Fig. 1.</a>).</li>
</ol>
</ol>
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</p><img src="https://static.igem.org/mediawiki/2008/e/ec/Blot_OmpA_WAW.jpg" width=150/>
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</p><a name="fig1"><img src="https://static.igem.org/mediawiki/2008/e/ec/Blot_OmpA_WAW.jpg" width=150/></a>
<var><b>Fig. 1.</b>Protein A detection on the surface of bacteria.<br>
<var><b>Fig. 1.</b>Protein A detection on the surface of bacteria.<br>
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1 - <i>E. coli</i> expressing <b>OmpA_Omega_A_Alpha</b>,<br>
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1 - <i>E. coli</i> expressing OmpA_Omega_A_Alpha,<br>
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2 - <b>negative control</b> - <i>E. coli</i> expressing <b>OmpA_Z_Omega</b>,<br>
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2 - <i>E. coli</i> expressing OmpA_Z_Omega (antibodies binds to protein Z probably due to similarity of protein A and Z),<br>
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3 - <i>E. coli</i> expressing <b>OmpA_A_Alpha</b>,<br>
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3 - <i>E. coli</i> expressing OmpA_A_Alpha,<br>
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4 - <b>negative control</b> - <i>E. coli</i> carrying OmpA_Omega_A_Alpha without inducer,<br>
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4 - <i>E. coli</i> carrying OmpA_Omega_A_Alpha without inducer,<br>
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5 - <i>E. coli</i> expressing <b>OmpA_A_Omega</b>,<br>
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5 - <i>E. coli</i> expressing OmpA_A_Omega,<br>
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6 - <b>negative control</b> - <i>E. coli</i> Top10 without plasmid.</var>
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6 - <i>E. coli</i> Top10 without plasmid.</var>
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<br><br>

Latest revision as of 18:04, 27 October 2008

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Checking for presence of protein A on the cell membrane

Piotr

  1. Pouring bacteria by drops onto nitrocellulose.
  2. Drying.
  3. Blocking.
  4. Anti-A antibody binding.
  5. Washing.
  6. Anti-rabbit antibody binding.
  7. Developing with BCIP and NBT (see Fig. 1.).

Fig. 1.Protein A detection on the surface of bacteria.
1 - E. coli expressing OmpA_Omega_A_Alpha,
2 - E. coli expressing OmpA_Z_Omega (antibodies binds to protein Z probably due to similarity of protein A and Z),
3 - E. coli expressing OmpA_A_Alpha,
4 - E. coli carrying OmpA_Omega_A_Alpha without inducer,
5 - E. coli expressing OmpA_A_Omega,
6 - E. coli Top10 without plasmid.

Cloning of alpha to pACYC177+OmpA_A_omega and pACYC177+OmpA_Z_omega

Michał K.

Inoculation of pACYC177+OmpA_A_omega and pACYC177+OmpA_Z_omega plasmids to LB with kanamycin.

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