User:Jorge E. Buendia Buendia/Notebook/iGEM UNAM-Genomics-Mexico/2010/08/02

{| width="800"
 * style="background-color: #EEE"|[[Image:igem-logo-150px.png|150px]] iGEM UNAM-Genomics-Mexico
 * style="background-color: #F2F2F2" align="center"|  |Main project page
 * style="background-color: #F2F2F2" align="center"|  |Main project page


 * colspan="2"|
 * colspan="2"|

August 2nd, 2010
1. Verify constructions by Colony PCR to amplify inserts from the following ligations:
 * L1: p30-MinBP + GFP E0240
 * L2: p30-MinBP + GFP BBa_K145015 (74 min)
 * L4: Backbone plasmid pSB3K3 + Blue Promoter BBa_K238013 + GFP BBa_K145015 (74 min)
 * L5: Plasmid 18 pSB1T3 + GFP BBa_K145015
 * L6: Religation p30-MinBP to make it a standard BioBrick


 * Primers used were Preffix FWD and Suffix REV, reactives needed for one reactions are as follows:
 * PCR with Taq DNA polymerase
 * '''Reactives (ul x sample)
 * Taq Polymerase -> 1
 * Taq Reaction Buffer 10X -> 5
 * MgCl 50mM (can be used up to 3ul) -> 2.5
 * dNTP’s 0.4ug/ul	-> 2.5
 * Primer Forward (can be used up to 3ul)	-> 2.5
 * Primer Reverse (can be used up to 3ul)	-> 2.5
 * HPLC -> 24
 * DNA -> 10
 * Total volume -> 50


 * Thermocycler program:
 * 1. 95ºC 5 min
 * 2. 30 cycles
 * 95ºC 45 seg
 * 60ºC 45 seg
 * 72ºC 1.5 min
 * 3. 72ºC 5 min
 * 4. Hold 4ºC

2. Run gel to verify Colony PCR.



Lanes: 1) Green ladder; 2) L1: p30-MinBP + GFP E0240; 3) L2: p30-MinBP + GFP BBa_K145015 (74 min); 4) L4: Backbone plasmid pSB3K3 + Blue Promoter BBa_K238013 + GFP BBa_K145015 (74 min); 5) L5: Plasmid 18 pSB1T3 + GFP BBa_K145015; 6) L6: Religation p30-MinBP to make it a standard BioBrick; 7) BBa_I20260 pSB3K3 + J23101 + GFP E0040


 * }