IGEM:British Columbia/2009/Notebook/Biosensor Sensitivity/2009/08/13

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assembled assemblies 2, 11, 12, 13, 27, 28, 22, 23, 24, 25
1. Digestion using standard UBC iGEM digestion protocol


 * Prefix samples (cut with EcoRI and SpeI)
 * BBa_J23100
 * Lock 1 (Amp)
 * Key 1 (Amp)
 * ASEM 21


 * Suffix samples (cut with XbaI and PstI)
 * ASEM 17
 * ASEM 18
 * ASEM 19
 * ASEM 20
 * BBa_B0014
 * BBa_E0022
 * BBa_E0034
 * BBa_J31001
 * BBa_J61046

pSB1C3 pSB1A3
 * Backbone cut (cut with PstI and EcoRI)

2. Ligation using standard UBC iGEM digestion protocol

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 * ligation into pSB1C3':
 * i) inserts
 * ASEM 02 (1.00μL)
 * ASEM 11 (7.09μL)
 * ASEM 12 (9.32μL)
 * ASEM 13 (1.10μL)
 * ASEM 27 (7.89μL)
 * ASEM 28 (8.83μL)
 * ii) reaction mixture
 * vector: 2μL
 * insert: please see volume in parathesis above
 * water: 14μL - insert volume
 * 10x buffer: 2μL
 * ligase: 1μL


 * ligation into pSB1A3':
 * i) inserts
 * ASEM 22 (10.42μL)
 * ASEM 23 (9.51μL)
 * ASEM 24 (9.94μL)
 * ASEM 25 (10.41μL)
 * ii) reaction mixture
 * vector: 2μL
 * insert: please see volume in parathesis above
 * water: 14μL - insert volume
 * 10x buffer: 2μL
 * ligase: 1μL

Overnight ligation in cold (ice) water bath

transformed 10μL of each ligation mix


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