Team:Stockholm/6 October 2010

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(New page: {{Stockholm/Top2}} ==Andreas==)
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{{Stockholm/Top2}}
==Andreas==
==Andreas==
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===Sequencing results===
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Received sequencing results for the following samples ([[media:Sequencings_fasta_6oct.txt|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)
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*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)
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*pEX.nTra10⋅SOD⋅His (pEXf)
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*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===
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''Of stored pellets and ON cultures from 5/10''
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 +
*E.Z.N.A. Plasmid Miniprep kit
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**50 μ elution volume (elution buffer)
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{|border="1" cellpadding="1" cellspacing="0"
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!colspan="3"|DNA concentration
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|-
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!Sample
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!width="60"|Conc [ng/μl]
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!width="60"|A<sub>260</sub>/A<sub>280</sub>
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|-
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|pEX.nTAT&sdot;SOD&sdot;His.RBS.yCCS
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|align="center"|107.5
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|align="center"|1.96
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|-
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|pEX.nTra10&sdot;SOD&sdot;His.RBS.yCCS
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|align="center"|108.7
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|align="center"|1.93
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|-
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|pEX.nLMWP&sdot;SOD&sdot;His.RBS.yCCS
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|align="center"|84.35
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|align="center"|1.97
 +
|-
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|pSB1C3.His&sdot;SOD&sdot;cLMWP B
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|align="center"|255.4
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|align="center"|1.94
 +
|-
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|pSB1C3.His&sdot;SOD&sdot;cLMWP C
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|align="center"|201.2
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|align="center"|1.94
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|-
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|pSB1C3.His&sdot;SOD&sdot;cTAT D
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|align="center"|369.5
 +
|align="center"|1.92
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|-
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|pSB1C3.His&sdot;SOD&sdot;cTra10 A
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|align="center"|188.6
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|align="center"|1.95
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|}
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 +
===BL21 transformation===
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Quick transformation w/ 30 min on ice, 50 sec heat-shock.
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*30 &mu;l competent BL21(DE3)
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*1 &mu;l plasmid
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**pEX.nTra10&sdot;SOD&sdot;His.RBS.yCCS
 +
**pEX.nTAT&sdot;SOD&sdot;His.RBS.yCCS
 +
**pEX.nLMWP&sdot;SOD&sdot;His.RBS.yCCS

Revision as of 16:40, 6 October 2010


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