Team:Stockholm/6 October 2010

From 2010.igem.org

(Difference between revisions)
(New page: {{Stockholm/Top2}} ==Andreas==)
(→Andreas)
Line 1: Line 1:
{{Stockholm/Top2}}
{{Stockholm/Top2}}
==Andreas==
==Andreas==
 +
 +
===Sequencing results===
 +
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)
 +
*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)
 +
 +
{|border="1" cellpadding="1" cellspacing="0"
 +
!colspan="3"|DNA concentration
 +
|-
 +
!Sample
 +
!width="60"|Conc [ng/μl]
 +
!width="60"|A<sub>260</sub>/A<sub>280</sub>
 +
|-
 +
|pEX.nTAT&sdot;SOD&sdot;His.RBS.yCCS
 +
|align="center"|107.5
 +
|align="center"|1.96
 +
|-
 +
|pEX.nTra10&sdot;SOD&sdot;His.RBS.yCCS
 +
|align="center"|108.7
 +
|align="center"|1.93
 +
|-
 +
|pEX.nLMWP&sdot;SOD&sdot;His.RBS.yCCS
 +
|align="center"|84.35
 +
|align="center"|1.97
 +
|-
 +
|pSB1C3.His&sdot;SOD&sdot;cLMWP B
 +
|align="center"|255.4
 +
|align="center"|1.94
 +
|-
 +
|pSB1C3.His&sdot;SOD&sdot;cLMWP C
 +
|align="center"|201.2
 +
|align="center"|1.94
 +
|-
 +
|pSB1C3.His&sdot;SOD&sdot;cTAT D
 +
|align="center"|369.5
 +
|align="center"|1.92
 +
|-
 +
|pSB1C3.His&sdot;SOD&sdot;cTra10 A
 +
|align="center"|188.6
 +
|align="center"|1.95
 +
|}
 +
 +
===BL21 transformation===
 +
Quick transformation w/ 30 min on ice, 50 sec heat-shock.
 +
*30 &mu;l competent BL21(DE3)
 +
*1 &mu;l plasmid
 +
**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

Home Project Idea Lab Work Results Modelling Team       Follow us on and        2010.igem.org

↳   Notebook Safety Protocols


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
Retrieved from "http://2010.igem.org/Team:Stockholm/6_October_2010"