Team:Newcastle/13 July 2010

From 2010.igem.org

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(Materials and Protocol)
(Protocol)
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==Protocol==
==Protocol==
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*''E. coli'' DH5alpha was [https://2010.igem.org/TeamNewcastleTransformation_of_E._coli#Protocol|transformed]] again with our ligation mix.
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*''E. coli'' DH5alpha was [[https://2010.igem.org/TeamNewcastleTransformation_of_E._coli#Protocol|transformed]] again with our ligation mix.
*The seven overnight cultures underwent [[Team:Newcastle/Qiagen Minipreps|plasmid extraction]].
*The seven overnight cultures underwent [[Team:Newcastle/Qiagen Minipreps|plasmid extraction]].
*The seven plasmid extracts were then [https://2010.igem.org/TeamNewcastleRestriction_digests| restriction digested] with ecoR1 to linearise.
*The seven plasmid extracts were then [https://2010.igem.org/TeamNewcastleRestriction_digests| restriction digested] with ecoR1 to linearise.

Revision as of 09:15, 10 August 2010

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Contents

LacI BioBrick Construction

Aims

To use PCR to extract lacI (promoter, ribosome-binding site (RBS) & coding sequence (CDS)) from plasmid pMutin4 and ligate into vector pSB1AT3 in front of red fluorescent protein (RFP).

Materials

Seven overnight cultures Restriction enzyme ecoR1

Protocol

  • E. coli DH5alpha was [[1]] again with our ligation mix.
  • The seven overnight cultures underwent plasmid extraction.
  • The seven plasmid extracts were then restriction digested with ecoR1 to linearise.
  • The seven digests were then run on a gel.

Results

This procedure was followed in order to determine whether our lacI had been ligated into the plasmid vector. The Gel shows that

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