Team:Stockholm/6 October 2010

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Contents

Andreas

Sequencing results

Received sequencing results for the following samples (fasta):

  • pSB1K3.nTra10⋅SOD⋅His.RBS.yCCS 1 (VF2 & VR)
  • pSB1K3.nTra10⋅SOD⋅His.RBS.yCCS 2 (VF2 & VR)
  • pSB1K3.nTAT⋅SOD⋅His.RBS.yCCS 2 (VF2 & VR)
  • pSB1K3.nTAT⋅SOD⋅His.RBS.yCCS 3 (VF2 & VR)
  • pSB1K3.nLMWP⋅SOD⋅His.RBS.yCCS 2 (VF2 & VR)
  • pSB1K3.nLMWP⋅SOD⋅His.RBS.yCCS 3 (VF2 & VR)
  • pEX.nTra10⋅SOD⋅His (pEXf)
  • pEX.nLMWP⋅SOD⋅His (pEXf)

Sequences aligned using Geneious software, showing correct sequences for all constructs. Operon constructs all had an insertion between SpeI and PstI; this will not affect expression.

Plasmid prep

Of stored pellets and ON cultures from 5/10

  • E.Z.N.A. Plasmid Miniprep kit
    • 50 μ elution volume (elution buffer)
DNA concentration
Sample Conc [ng/μl] A260/A280
pEX.nTAT⋅SOD⋅His.RBS.yCCS 107.5 1.96
pEX.nTra10⋅SOD⋅His.RBS.yCCS 108.7 1.93
pEX.nLMWP⋅SOD⋅His.RBS.yCCS 84.35 1.97
pSB1C3.His⋅SOD⋅cLMWP B 255.4 1.94
pSB1C3.His⋅SOD⋅cLMWP C 201.2 1.94
pSB1C3.His⋅SOD⋅cTAT D 369.5 1.92
pSB1C3.His⋅SOD⋅cTra10 A 188.6 1.95

BL21 transformation

Quick transformation w/ 30 min on ice, 50 sec heat-shock.

  • 30 μl competent BL21(DE3)
  • 1 μl plasmid
    • pEX.nTra10⋅SOD⋅His.RBS.yCCS
    • pEX.nTAT⋅SOD⋅His.RBS.yCCS
    • pEX.nLMWP⋅SOD⋅His.RBS.yCCS