IGEM:Harvard/2009/Notebook/Harvard iGEM 2010/2010/08/02: Difference between revisions
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[[Image:Gal4_Ecr_digest_gel_ex_8-2.jpg]] | [[Image:Gal4_Ecr_digest_gel_ex_8-2.jpg]] | ||
===Nanodrop of Gal4 and EcR Gel Extractions=== | |||
{| border="3" style="margin-left: 3em;" | |||
|- | |||
! Plasmid !! Quantity (ng/μL) !! 260/280 | |||
|- | |||
! Gal4 (1) | |||
| 1.4 || -3.18 | |||
|- | |||
! Gal4 (2) | |||
| 7.2 || 4.59 | |||
|- | |||
! EcR (1) | |||
| 5.3 || 2.52 | |||
|- | |||
! EcR (2) | |||
| 5.6 || 2.12 | |||
|} | |||
===VP16 PCR 2=== | ===VP16 PCR 2=== | ||
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[[Image:VP16PCR.2_blank_8-2.jpg]] | [[Image:VP16PCR.2_blank_8-2.jpg]] | ||
===VP16 PCR.3=== | |||
60μL reaction: | |||
*1.5 μL pfx polymerase | |||
*3μL DNTPs | |||
*18μL enhancer buffer | |||
*1.5μL MgSO<sub>4</sub> | |||
*3μL 1/10 dilution VP16.Hind.Fwd | |||
*3μL 1/10 dilution VP16.Rev | |||
*3μL 1/10 dilution of VP16 (1) | |||
*12μL amp buffer | |||
*5μL DH<sub>2</sub>O | |||
Using modified program: | |||
1=94°C for 2:00 | |||
2=94°C for :15 | |||
3=60°C for :30 | |||
4=70°C for :45 | |||
5=Goto 2, 30 times | |||
6=4.0°C forever | |||
7=end | |||
(see tomorrow's entry for gel image) | |||
===Ligations=== | ===Ligations=== |
Revision as of 13:43, 7 September 2010
iGEM iGarden | <html><img src="/images/9/94/Report.png" border="0" /></html> Main project page <html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>Previous entry<html> </html>Next entry<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html> | |||||||||||||||
Team FenceGel extractionGal4 spe bam 8/2 and EcR bam xba 8/2 gel extracted and purified. Gal4 on the left, EcR on the right Nanodrop of Gal4 and EcR Gel Extractions
VP16 PCR 260μL reaction:
using program: 1=94°C for 2:00 2=94°C for :15 3=68°C for :30 4=Goto 2, 30 times 5=4.0°C forever 6=end
VP16 PCR.360μL reaction:
Using modified program: 1=94°C for 2:00 2=94°C for :15 3=60°C for :30 4=70°C for :45 5=Goto 2, 30 times 6=4.0°C forever 7=end (see tomorrow's entry for gel image) LigationsGal4 into EcR Barnase into B15 LTP into B11 (positive control from team allergy) Gal4
EcR control
Barnase
B15 control
LTP
B11 control
Allowed tubes to sit at room temp for an hour, proceeded to transformation Team FlavorDTL of Mira/Brazz+StrepII+Stop into V24
DTL of Wintergreen pathway
Gel Purification
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