Team:Warsaw/Calendar-Main/23 June 2008

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<h3> Blue/white and rifampicin test #2 </h3>
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<h4>Piotr</h4>
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<p>Analysis of lacZ sequence from <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pZC320>pZC320</a>:<br>
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There is no mutation in lacZ gene. Why are the colonies white? Reason of white phenotype unknown; this might be chromosomal mutation, transcription supression or something else). </p>
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<h3>Preparation of constructs with OmpA protein fusions<br>
 
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Michał K.</h3>
 
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<li> Inoculation other separate transformant colonies from plates with ligations: pACYC177+OmpA_alpha pACYC177+OmpA_omega (liquid LB + kanamycin).</li>
 
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<h3>PCR results<br>
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<h3>Preparation of constructs: OmpA_alpha and OmpA_omega</h3>
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Michał L., Ewa, Marcin</h3>
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<h4>Michał K.</h4>
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<p>We have successfully amplified Omega-linker, but were unable to obtain the other two. PCRs were repeated. New annealing temp.: 48&deg;C. Mg<sup>2+</sup> concentration elevated to 12 μM.</p>
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<p>
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Inoculation other separate transformant colonies from plates with ligations: <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-alpha>pACYC177+OmpA_alpha </a> and <a href=https://2008.igem.org/Wiki/Team:Warsaw/vectors/pACYC177%2BompA-omega>pACYC177+OmpA_omega</a> (liquid LB + kanamycin).</p>
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<h3>Preparation of alpha-A and omega-A fusions</h3>
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<h4>Michał L.</h4>
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<p>We have successfully amplified Omega-linker (<a href="https://2008.igem.org/Team:Warsaw/Calendar-Main/23_June_2008#fig1">Fig. 1</a>), but were unable to obtain the other. two PCRs were repeated. New annealing temp.: 48&deg;C. Mg<sup>2+</sup> concentration elevated to 4 mM.</p>
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<a name="fig1"><img src="https://static.igem.org/mediawiki/2008/3/36/PCR_Alinker_Omegalinker_WAW.jpg"width=200/></a>
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<var><b>Fig. 1.</b>PCR products. Lane 1 - lack of product for A-linker; lane 3 - product for Omega-linker.</var>
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Blue/white and rifampicin test #2

Piotr

Analysis of lacZ sequence from pZC320:
There is no mutation in lacZ gene. Why are the colonies white? Reason of white phenotype unknown; this might be chromosomal mutation, transcription supression or something else).

Preparation of constructs: OmpA_alpha and OmpA_omega

Michał K.

Inoculation other separate transformant colonies from plates with ligations: pACYC177+OmpA_alpha and pACYC177+OmpA_omega (liquid LB + kanamycin).

Preparation of alpha-A and omega-A fusions

Michał L.

We have successfully amplified Omega-linker (Fig. 1), but were unable to obtain the other. two PCRs were repeated. New annealing temp.: 48°C. Mg2+ concentration elevated to 4 mM.

Fig. 1.PCR products. Lane 1 - lack of product for A-linker; lane 3 - product for Omega-linker.

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