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IGEM:University of Illinois Urbana Champaign/2009/Notebook/Bioware 2010 Gold Bioremediation/2010/10/26

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10.26.10

Minipreps were made from:

  • goLS + pSBIC3
  • golB + pSBIC3
  • golT + pSBIC3
  • golS + RBS + Terminator + pSBIC3
  • golB + RBS + Terminator + pSBIC3
  • golT + RBS + Terminator + pSBIC3

The plasmids that only had the single part were put into the cryostock and labeled:

  • ISB 240 - golB on pSBIC3
  • ISB 241 - golS on pSBIC3
  • ISB 242 - golS + B0034 + B0015 + pSBIC3
  • ISB 243 - golB + B0034 + B0015 + pSBIC3

Digestions were ran for the following:

  • golS + RBS + Terminator
  • golB + RBS + Terminator
  • golT + RBS + Terminator
  • J23119

Sequencing of; golS + RBS + Terminator, golB + RBS + Terminator, golT + RBS + Terminator was done using the VF2 primer to verify.

A gel for the earlier digestions was done and parts were extracted from the gel.

Ligations using the extracted genes from the gel:

  • golS + RBS + Terminator + J23119 + pSBIT3
  • golB + RBS + Terminator + J23119 + pSBIT3
  • golT + RBS + Terminator + J23119 + pSBIT3

After the ligations, PCR was used to verify that the inserts were in the plasmid backbone using VF2/VR primers. The ligations were put into cells via heat shock transformation.