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Team:Stockholm/6 October 2010
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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
