Team:TU Delft/Project/rbs-characterization/parts

From 2010.igem.org

(Difference between revisions)
(New page: =BioBricks, the making of= BBa_I13401 was PCR amplified using the universal primers G00100 and G00101. The purified PCR product was assembled into one of the Anderson RBS plasmids provided...)
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=BioBricks, the making of=
=BioBricks, the making of=
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BBa_I13401 was PCR amplified using the universal primers G00100 and G00101. The purified PCR product was assembled into one of the Anderson RBS plasmids provided in the distribution plates by means of 2-way ligations thus obtaining:
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For our RBS characterization project, five different RBS sequences from the [http://partsregistry.org/Ribosome_Binding_Sites/Prokaryotic/Constitutive/Anderson Anderson RBS family] (J61100, J61101, J61107, J61117, J61127) and the standard RBS [http://partsregistry.org/Part:BBa_B0032 B0032] were placed in front of the standard [http://partsregistry.org/Part:BBa_E0040 GFP coding sequence].
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The Biobricks generated in order to perform the experiments were: [http://partsregistry.org/Part:BBa_K398500 K398500], [http://partsregistry.org/Part:BBa_K398501 K398501], [http://partsregistry.org/Part:BBa_K398502 K398502], [http://partsregistry.org/Part:BBa_K398503 K398503], [http://partsregistry.org/Part:BBa_K398504 K398504]. The general map of the construction is shown below, where the RBS is displayed in fucsia.
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*K398500 (BBa_J61100 + BBa_I13401)
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[[Image:RBS1.jpg|left|650px]]
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*K398501 (BBa_J61101 + BBa_I13401)
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*K398502 (BBa_J61107 + BBa_I13401)
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{| style="color:black; background-color:white;" cellpadding="5" cellspacing="0" border="1"
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*K398503 (BBa_J61117 + BBa_I13401)
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|'''Feature'''
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*K398504 (BBa_J61127 + BBa_I13401)
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|'''Function'''
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|-
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|[http://partsregistry.org/Part:BBa_B0032 AmpR]
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|Ampicillin resistance
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|-
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|[http://partsregistry.org/Part:BBa_B0015 B0015]
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|Transcriptional (double) terminator
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|-
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|[http://partsregistry.org/Part:BBa_B0062 B0062]
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|Transcriptional terminator
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|-
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|[http://partsregistry.org/Part:BBa_E0040 E0040]
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|GFP
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|-
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|[http://partsregistry.org/Part:BBa_G00000 G00000]
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|Standard prefix
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|-
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|[http://partsregistry.org/Part:BBa_G00001 G00001]
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|Standard suffix
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|-
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|[http://partsregistry.org/Part:BBa_G00100 G00100]
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|VF2 primer binding site
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|-
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|[http://partsregistry.org/Part:BBa_G00101 G00101]
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|VR primer binding site
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|-
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|[http://partsregistry.org/Part:BBa_J61100 J61100]
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|RBS Anderson family
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|-
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|[http://partsregistry.org/Part:BBa_J23100 J23100]
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|Promoter
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|}
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BBa_I13401 was PCR amplified using the universal primers G00100 and G00101. The purified PCR product was assembled into one of the Anderson RBS plasmids provided in the distribution plates by means of 2-way ligations.
Transformation into competent Top10 E.coli cells yielded positives, as determined by fluorescence analysis.
Transformation into competent Top10 E.coli cells yielded positives, as determined by fluorescence analysis.

Revision as of 16:55, 10 September 2010

BioBricks, the making of

For our RBS characterization project, five different RBS sequences from the [http://partsregistry.org/Ribosome_Binding_Sites/Prokaryotic/Constitutive/Anderson Anderson RBS family] (J61100, J61101, J61107, J61117, J61127) and the standard RBS [http://partsregistry.org/Part:BBa_B0032 B0032] were placed in front of the standard [http://partsregistry.org/Part:BBa_E0040 GFP coding sequence]. The Biobricks generated in order to perform the experiments were: [http://partsregistry.org/Part:BBa_K398500 K398500], [http://partsregistry.org/Part:BBa_K398501 K398501], [http://partsregistry.org/Part:BBa_K398502 K398502], [http://partsregistry.org/Part:BBa_K398503 K398503], [http://partsregistry.org/Part:BBa_K398504 K398504]. The general map of the construction is shown below, where the RBS is displayed in fucsia.

RBS1.jpg
Feature Function
[http://partsregistry.org/Part:BBa_B0032 AmpR] Ampicillin resistance
[http://partsregistry.org/Part:BBa_B0015 B0015] Transcriptional (double) terminator
[http://partsregistry.org/Part:BBa_B0062 B0062] Transcriptional terminator
[http://partsregistry.org/Part:BBa_E0040 E0040] GFP
[http://partsregistry.org/Part:BBa_G00000 G00000] Standard prefix
[http://partsregistry.org/Part:BBa_G00001 G00001] Standard suffix
[http://partsregistry.org/Part:BBa_G00100 G00100] VF2 primer binding site
[http://partsregistry.org/Part:BBa_G00101 G00101] VR primer binding site
[http://partsregistry.org/Part:BBa_J61100 J61100] RBS Anderson family
[http://partsregistry.org/Part:BBa_J23100 J23100] Promoter

BBa_I13401 was PCR amplified using the universal primers G00100 and G00101. The purified PCR product was assembled into one of the Anderson RBS plasmids provided in the distribution plates by means of 2-way ligations.

Transformation into competent Top10 E.coli cells yielded positives, as determined by fluorescence analysis.

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