Team:Groningen/Notebook/22 August 2009

From 2009.igem.org

(Difference between revisions)
m (GVP Cluster)
m (GVP Cluster)
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:→ {{done}} Place the four test cultures with and without GVP on table at room temperature (make foto)
:→ {{done}} Place the four test cultures with and without GVP on table at room temperature (make foto)
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:→ {{todo}} Look at plates stored at 37C with BBa_J61002-pArsR<sup>+</sup>/pZntR<sup>+</sup>/pCueO<sup>+</sup> and use colonies to grow o.n. cultures for plasmid isolation
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:→ {{done}} Look at plates stored at 37C with BBa_J61002-pArsR<sup>+</sup>/pZntR<sup>+</sup>/pCueO<sup>+</sup> and use colonies to grow o.n. cultures for plasmid isolation
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'''Plasmid Purification'''
'''Plasmid Purification'''
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Plasmid isolation was performed on the cultures of E.coli TOP10 containing plasmids [http://partsregistry.org/wiki/index.php?title=Part:pSB3K3 pSB3K3] with [http://partsregistry.org/wiki/index.php?title=Part:BBa_J23100 high], [http://partsregistry.org/wiki/index.php?title=Part:BBa_J23106 medium] and [http://partsregistry.org/wiki/index.php?title=Part:BBa_J23109 low] constitutive promoters and GVP with the "Sygma-Aldrich™ [http://www.sigmaaldrich.com/life-science/molecular-biology/dna-and-rna-purification/plasmid-miniprep-kit.html GenElute™] Plasmid Miniprep Kit".
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Plasmid isolation was performed on the cultures of E.coli TOP10 containing plasmids [http://partsregistry.org/wiki/index.php?title=Part:BBa_J61002 BBa_J61002] with [http://partsregistry.org/wiki/index.php?title=Part:BBa_J23101 BBa_J23101] constitutive promoter with the "Sygma-Aldrich™ [http://www.sigmaaldrich.com/life-science/molecular-biology/dna-and-rna-purification/plasmid-miniprep-kit.html GenElute™] Plasmid Miniprep Kit".
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* From each tube 4mL of culture was collected in a 2.0mL cup, and the cells were pelleted by centrifugation for 1 min. at max. speed and the supernatant discarded.
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* From each tube 4mL of culture was collected in one 2.0mL cup, and the cells were pelleted by centrifugation for 1 min. at max. speed and the supernatant discarded.
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* Plasmids were eluted with 30μL MQ and stored in the fridge
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* Plasmids were eluted with 40μL MQ and stored in the fridge
===Transporters===
===Transporters===

Revision as of 09:24, 22 August 2009

Igemhomelogo.png

Wet

GVP Cluster

DONE Isolate plasmids from E.coli Top10 BBa_J23101 cells
DONE Plate E.coli Top10 BBa_J23101 cells from o.n. culture
TODO Pellet remaining cells for short storage in -20 box (not needed)
DONE Place the four test cultures with and without GVP on table at room temperature (make foto)
DONE Look at plates stored at 37C with BBa_J61002-pArsR+/pZntR+/pCueO+ and use colonies to grow o.n. cultures for plasmid isolation


Plates

Showed single colony growth on plates with J61002-pMEtal+RBS-RFP plasmids, and were stored in the fridge for future preculture growth.

→ The plates with high and low concentration of transformed cells showed colonies in the expected ratio.
→ On the plates for pZntR+ and pCueO+ no colonies were dark red, indication of ligation of the original high constitutive promoter back into the J61002 vector, or uncut plasmid which was transformed.
→ The plates with pArsR+ plasmid showed only a few non-coloured colonies, mostly medium red colour, andn a few dark red colonies. It was already thought that the pArsR is a bit leaky and causes expression of RFP.

Over Night Cultures

→ Both cultures of BBa_J61002-J23101 showed growth, and can be used for plasmid isolation.
www.sigmaaldrich.com

Plasmid Purification

Plasmid isolation was performed on the cultures of E.coli TOP10 containing plasmids BBa_J61002 with BBa_J23101 constitutive promoter with the "Sygma-Aldrich™ GenElute™ Plasmid Miniprep Kit".

  • From each tube 4mL of culture was collected in one 2.0mL cup, and the cells were pelleted by centrifugation for 1 min. at max. speed and the supernatant discarded.
  • Plasmids were eluted with 40μL MQ and stored in the fridge

Transporters

Metal Accumulation

Vectors

Dry

April
MTWTFSS
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May
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25 26 27 28 29 30 31
June
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15 16 17 18 19 20 21
22 23 24 25 26 27 28
29 30
July
MTWTFSS
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13 14 15 16 17 18 19
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27 28 29 30 31
August
MTWTFSS
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10 11 12 13 14 15 16
17 18 19 20 21 22 23
24 25 26 27 28 29 30
31
September
MTWTFSS
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7 8 9 10 11 12 13
14 15 16 17 18 19 20
21 22 23 24 25 26 27
28 29 30
October
MTWTFSS
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5 6 7 8 9 10 11
12 13 14 15 16 17 18
19 20 21 22 23 24 25
26 27 28 29 30 31
November
MTWTFSS
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9 10 11 12 13 14 15
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30