IGEM:Arizona State/2014/Notebook/asu igem 2014/2014/07/15

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(Autocreate 2014/07/15 Entry for IGEM:Arizona_State/2014/Notebook/asu_igem_2014)
Current revision (10:46, 16 July 2014) (view source)
(TesA/RBS Ligation Attempt 2)
 
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==Entry title==
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==TesA/RBS Ligation Attempt 2==
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* Insert content here...
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* Checked DNA concentrations
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:TesA= 11.99 ng/μL ; RBS=32.99 ng/μL
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*When divided by the number of base pairs: [TesA]=.018 and [RBS]=.016
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* Ligation
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{| {{table}} border="1" align=center <!-- Ligation table -->
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|- valign="top"
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| bgcolor=#cfcfcf | Reagent (μL)
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| bgcolor=#cfcfcf | Volume (μL)
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|-
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|| DNA(plasmid) || 1.0 μL RBS, 2.0 μL TesA
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|-
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|| 10X buffer || 1.0
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|-
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|| dH<sub>2</sub>O || 5.0
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|-
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|| Ligaze || 1.0
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|-
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|| &nbsp; || 10.0 μL --> 25°C/ ~1 Hr.
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|}
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*After ligation, the mixture was heated to 65°C for 10 min. to denature the ligaze.
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*Transformation was then performed.
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:Three plates were grown overnight: RBS backbone (vector), TesA+RBS, and mCherryboth as a positive control and to replenish our stocks.
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* Transformation
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:Unfortunately, no growth was seen on any of the plates, including the positive control of mCherry.
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TesA/RBS Ligation Attempt 2

  • Checked DNA concentrations
TesA= 11.99 ng/μL ; RBS=32.99 ng/μL
  • When divided by the number of base pairs: [TesA]=.018 and [RBS]=.016
  • Ligation
Reagent (μL) Volume (μL)
DNA(plasmid) 1.0 μL RBS, 2.0 μL TesA
10X buffer 1.0
dH2O 5.0
Ligaze 1.0
  10.0 μL --> 25°C/ ~1 Hr.
  • After ligation, the mixture was heated to 65°C for 10 min. to denature the ligaze.
  • Transformation was then performed.
Three plates were grown overnight: RBS backbone (vector), TesA+RBS, and mCherryboth as a positive control and to replenish our stocks.
  • Transformation
Unfortunately, no growth was seen on any of the plates, including the positive control of mCherry.
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