Team:DTU-Denmark/Notebook/30 August 2013

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===PCR to amplify HAO and cycAX USER fragments===
===PCR to amplify HAO and cycAX USER fragments===
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Made 6 reactions for each, using either the extraction PCR fragment or the glycerol stock as template
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{| class="wikitable" style="text-align: right"
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! label !! cyc gly D !! cyc gly % !! cyc gly 1uL D !! cyc ext D !! cyc ext % !! cyc ext 1uL D !!
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|-
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|  template || glycerol stock || glycerol stock || glycerol stock || extraction PCR fragment || extraction PCR fragment || extraction PCR fragment
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|-
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|  DMSO || 1,5 || - || 1,5 || 1,5 || - || 1,5
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|-
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|  MgCl2 || - || - || 1 || - || - || 1
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|-
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|  MQ || 1 || 2,5 || - || 1 || 2,5 || -
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|-
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|}
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Program:
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{| class="wikitable" style="text-align: right"
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! temperature !! time !! cycles
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|-
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|  98C || 2:00 || -
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|-
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|  98C || 0:20 || 36
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|-
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|  annealing temperature || 1:00 || 36
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|-
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|  72C || extension time || 36
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|-
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|  72C || 5:00 || -
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|-
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|  10C || hold || -
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|-
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|}
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HAO - 59C, 3:00
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cycAX - 55C, 1:30
==Results==
==Results==

Revision as of 17:37, 30 August 2013

30 August 2013

Contents

lab 208

Main purpose

Who was in the lab

Procedure

PCR to amplify HAO and cycAX USER fragments

Made 6 reactions for each, using either the extraction PCR fragment or the glycerol stock as template

label cyc gly D cyc gly % cyc gly 1uL D cyc ext D cyc ext % cyc ext 1uL D
template glycerol stock glycerol stock glycerol stock extraction PCR fragment extraction PCR fragment extraction PCR fragment
DMSO 1,5 - 1,5 1,5 - 1,5
MgCl2 - - 1 - - 1
MQ 1 2,5 - 1 2,5 -


Program:

temperature time cycles
98C 2:00 -
98C 0:20 36
annealing temperature 1:00 36
72C extension time 36
72C 5:00 -
10C hold -

HAO - 59C, 3:00 cycAX - 55C, 1:30

Results

Gel

  • 1 kb ladder
  • psB1C3 for Nir, 2% DMSO
  • psB1C3 for Nir, 2% DMSO, 1 uL 50mM MgCl2
  • psB1C3 for Nir, 4% DMSO
  • psB1C3 for Nir, 4% DMSO, 1 uL 50mM MgCl2
  • psB1C3 for Nir, 1M Betaine
  • purification of psB1C3 USER fragment to make Biobricks (escaped from well -> Ethanol?)
  • purification of HAO fragment (escaped from well -> Ethanol?)
  • purification of AMO fragment (escaped from well -> Ethanol?)
  • purification of psB1C3 USER fragment to make Biobricks (used more loading buffer)
  • purification of HAO fragment (used more loading buffer)
  • purification of AMO fragment (used more loading buffer)
  • purification of pZA21::ara(tight)
  • purification of pZA21::ara(tight) with endings for Nir
  • 1 kb ladder

2013-08-30 psbnir puri verification2.jpg

Conclusion

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