Team:NTU-Singapore/Notebook/11 July 2008

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(New page: *SupD gel check showed that PCR product was not SupD - re-ran the PCR and made sure the product was right this time. *GFP-6 and E7 were digested. *Gel checks were also ran for T7ptag & E...)
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Revision as of 18:58, 14 July 2008

  • SupD gel check showed that PCR product was not SupD - re-ran the PCR and made sure the product was right this time.
  • GFP-6 and E7 were digested.
  • Gel checks were also ran for T7ptag & E7. Darius found out that running SupD on a 1.5% agarose gel, compared to the usual 1% gel would yield clearer bands - this proved to be true.
  • Purification problems..

..were solved with the addition of 10μl of Proteinase K to 50μl of PCR product, then incubated at 37degC for 30mins, then 68degC for a further 10 minutes. This yielded clear bands that were otherwise smeary and thus indistinguishable.

  • Gel extraction of E7 & T7ptag (E-P) was also carried out.
    • GFP (X-P) & E7 (X-P) produced no bands.
  • Gel run for LacI-GFP & Fe-GFP
  • Gel check for re-digested Fe.
  • Ligation
    • Two protocols were carried out, one was Drew Endy's protocol (using T4 DNA Ligase) and the other was Quick Ligase, both on E7 and T7ptag.
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