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Team:Brown-Stanford/Lab/Protocols/Gibson
From 2011.igem.org
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{{:Team:Brown-Stanford/Templates/Protocol}} | {{:Team:Brown-Stanford/Templates/Protocol}} | ||
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== '''Gibson Assembly''' == | == '''Gibson Assembly''' == | ||
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| + | ''5x isothermal buffer''' | ||
*0.75g - 25% PEG-8000 | *0.75g - 25% PEG-8000 | ||
*1500 µL - 500 mM Tris-HCl pH 7.5 | *1500 µL - 500 mM Tris-HCl pH 7.5 | ||
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*216.75 µL - Nuclease-free water | *216.75 µL - Nuclease-free water | ||
'''Total - 375 µL''' | '''Total - 375 µL''' | ||
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{{:Team:Brown-Stanford/Templates/Foot}} | {{:Team:Brown-Stanford/Templates/Foot}} | ||
Revision as of 01:39, 10 July 2011
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REGObricks
Intro--------------------- ISRU--------------------- Biocement--------------------- S. pasteurii--------------------- Balloon
Flights--------------------- Transforming--------------------- Biobrick2 -
PowerCell
Intro--------------------- Cyanobacteria--------------------- Mars--------------------- Nutrients -
FRETcetera
-
Lab
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Partners
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Outreach
SB5.0--------------------- NASA--------------------- Lunar Science--------------------- Maker Faire--------------------- BBC--------------------- CBricks--------------------- Ethics--------------------- Regional
Jamboree
Code: A1
Gibson Assembly
5x isothermal buffer'
- 0.75g - 25% PEG-8000
- 1500 µL - 500 mM Tris-HCl pH 7.5
- 75 µL - 50mM MgCl2
- 150 µL - 50mM DTT
- 30 µL - 1mM dATP
- 30 µL - 1mM dTTP
- 30 µL - 1mM dCTP
- 30 µL - 1mM dGTP
- 300 µL - 5mM NAD
- Nuclease-free water to fill to 3000µL
Total - 3000 µL
Gibson Master Mix
- 50 µL - Taq ligase (40u/µL)
- 100 µL - 5x isothermal buffer
- 2 µL - T5 exonuclease (1u/µL)
- 6.25 µL - Phusion polymerase (2u/µL)
- 216.75 µL - Nuclease-free water
Total - 375 µL
