Team:Warsaw/Calendar-Main/24 September 2008

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  primers (20 cycles, elongation duration 15 s, annealing temperature 72&deg;C). </li>
  primers (20 cycles, elongation duration 15 s, annealing temperature 72&deg;C). </li>
<li> <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#pcr">PCR</a> on <a href=http://www.fermentas.com/techinfo/nucleicacids/mappuc1819.htm>pUC19</a> plasmid with  
<li> <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#pcr">PCR</a> on <a href=http://www.fermentas.com/techinfo/nucleicacids/mappuc1819.htm>pUC19</a> plasmid with  
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<a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#AlphaL+SacI">AlphaL+SacI</a> and <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#AlphaP_XB">AlphaP_XB</a>
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<a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#AlphaL+SacI">AlphaL+SacI</a> and <a href="https://2008.igem.org/Wiki/Team:Warsaw/primers#AlphaP+link10+homo2">AlphaP+link10+homo2</a>
  primers (20 cycles, elongation duration 45 s, annealing temperature 63&deg;C).
  primers (20 cycles, elongation duration 45 s, annealing temperature 63&deg;C).
<br>
<br>

Revision as of 22:31, 13 October 2008

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Mutagenesis of protein A

Paweł

  1. Results of mutagenesis: no colonies on any plate.
  2. Mutagenesis repeated with modified conditions: fresh stock of dNTPs used and additional MgCl2 added.

Preparation of alpha+A construct

Antoni

  1. PCR on pKS plasmid containing protein A with AL+link10+homo2 and AP+NotI primers (20 cycles, elongation duration 15 s, annealing temperature 72°C).
  2. PCR on pUC19 plasmid with AlphaL+SacI and AlphaP+link10+homo2 primers (20 cycles, elongation duration 45 s, annealing temperature 63°C).
    As a result we got two PCR products (alpha-linker and linker-A) wich will be utilized as templates in next PCR.
  3. Gel electrophoresis of PCR products and gel-out of proper bands (alpha_linker - 572 bp and linker_A - 467 bp ).
  4. Measurment of concentration of both isolated products

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