Team:Warsaw/Calendar-Main/2 October 2008

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<h3>Preparation of <a href=http://partsregistry.org/Part:pSB2K3>pSB2K3</a> + _alpha</h3>
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<h4>Michał K.</h4>
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<ol><li> Overnight <a href="https://2008.igem.org/wiki/index.php?title=Wiki/Team:Warsaw/protocols#ligation">ligation</a> of isolated DNA fragments: <a href=http://partsregistry.org/Part:pSB2K3>pSB2K3</a> + _alpha.</li>
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<h3>Preparation of BioBricks</h3>
<h3>Preparation of BioBricks</h3>

Revision as of 21:32, 26 October 2008

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Preparation of pSB2K3 + _alpha

Michał K.

  1. Overnight ligation of isolated DNA fragments: pSB2K3 + _alpha.

    2. Preparation of BioBricks

    Michał K.

    1. Isolation of plasmids from culture inoculated on previous day (pSB1A3+Z(BBa_K103004)).
    2. Control digest of isolated plasmid with EcoRI and PstI (Orange buffer). Gel electrophoresis - proper clones found.
    3. Digest of pLac_OmpA_omega fragment with EcoRI and BcuI (BamHI buffer).
    4. Gel elctrophoresis and gel-out of proper band - 1200 bp.
    5. Colony PCR with OmpaL_N and OmpaP_link primers on colonies from plates with transformations pSB1A3+OmpA (annealing temperature - 55°C,45 s of elongation step).
    6. Gel electrophoresis.
    7. Confirmed transformant colonies inoculated to liquid LB with ampicillin.
    8. Overnight ligation of isolated DNA fragments: pSB2K3 + _alpha, pSB2K3 + _omega and pACYC177 + OmpA_omega_ (both digested with BamHI and PstI - from 30 September).

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