User:Karmella Haynes/Notebook/BioBrick cloning/2009/12/08

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12/08/09 ✓ Assemblies: KAH109-116 Assemblies KAH109: (1) KAH87/(E/S)/1053 + (8) NLS-myc-stop(KAH011)/(E/X)/~3300 KAH110: (2) KAH88/(E/S)/1053 + " KAH111: (3) KAH89/(E/S)/1053 + " KAH112: (4) KAH90/(E/S)/1053 + " KAH113: (5) KAH91/(E/S)/2037 + " KAH114: (6) KAH92/(E/S)/2040 + " KAH115: (7) KAH74/(E/S)/861 + " KAH116: mCh/(E/S)/705 ✓ + " > Digests (Fermentas FD) Reagent Volume   30 μL/lane, 1% agarose DNA (plasmid) up to 25 μL 10x buffer 3.0 enzyme 1 1.0 enzyme 2 1.0 dH2O ---   30 μL --> 37°C/ ~30 min. > Measure conc.'s Sample OD260 260/280 ng/μL 1. KAH87(E/S) 0.176 2.45 8.8 2. KAH88(E/S) 0.202 2.01 10.1 3. KAH89(E/S) 0.128 2.76 6.4 4. KAH90(E/S) 0.136 2.44 6.8 5. KAH91(E/S) 0.151 2.28 7.5 6. KAH92(E/S) 0.108 2.50 5.4 7. KAH74(E/S) 0.101 3.24 5.1 8. KAH011(E/X) 0.863 2.00 43.2 > Ligations Ligation Plate results (lig : neg crtl) 12/09/09 1. KAH87(E/S)/1053, 13 ng + KAH011(E/X)/3300, 20 ng KAH109 >10:1 (Pick 2) 2. KAH88(E/S)/1053, 13 ng + " KAH110 >10:1 (Pick 2) 3. KAH89(E/S)/1053, 13 ng + " KAH110 >10:1 (Pick 2) 4. KAH90(E/S)/1053, 13 ng + " KAH110 >10:1 (Pick 2) 5. KAH91(E/S)/2037, 25 ng + " KAH110 >10:1 (Pick 2) 6. KAH92(E/S)/2040, 25 ng + " KAH110 >10:1 (Pick 2) 7. KAH74(E/S)/861, 10 ng + " KAH110 >10:1 (Pick 2) 8. mCh(E/S)/705, 9 ng + " KAH110 >10:1 (Pick 2) 9. KAH011(E/X) 20 ng     1 2 3 4 5 6 7 8 9 Insert DNA 1.5 1.3 2.0 1.9 3.3 4.5 2.0 1.1 --- Vector DNA 0.5 0.5 0.5 0.5 0.5 0.5 0.5 0.5 0.5 2x lgn buf (Roche) 6.0 6.0 6.0 6.0 6.0 6.0 6.0 6.0 6.0 T4 ligase (NEB) 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 dH2O 3.0 3.2 2.5 2.6 1.2 --- 2.5 3.4 4.5   12μL 12μL 12μL 12μL 12μL 12μL 12μL 12μL 12μL --> R.T./~10 min.; Add 20 μL Z-DH5α