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Team:DTU-Denmark/Notebook/26 June 2013
From 2013.igem.org
Contents |
208 lab
Main purposes today
helloworld-project
- 25 rxns including duplicates
- 34 rxna including duplicates with the following setup:
Tube 1-6 containing 5uL pZA21 plasmid template and 3uL of each primers for amplification of the backbone. Tube 7-10; 1uL g-Block from previous purification and 3uL of each Sec signal primers. Tube 11-14 containing same amount of g-Block and primers but now replaced with the TAT2 signal primers. Tube 15-18; 5uL GFP SF plasmid template and 3uL of each primers for GFP SF TAT. Tube 19-22; 5uL GFP SF plasmid template and 3uL of each primers for GFP SF Sec. Tube 23-26; 5uL RFP plasmid template and 3uL of each primer for RFP. Tube 27-30; 1uL g-Block purification and 3uL of each TAT3 primers. Tube 31-34; 0.5uL of the original g-Block and 3uL of each primers for amplifying the g-Block. All tube with less than 5uL template DNA was filled with MQ until final volume of 11uL(5+3+3).
First half of the samples where run on PCR with x7-polymerase and the second half where run with PHUSION-polymerase, with the exception of the g-Block where all reactions where run with PHUSION. This gives following setup: x7-poly in tube:
- 1-3, 7-8, 11-12, 15-16, 19-20, 23-24, 27-28
PHUSION in tube:
- 4-6, 9-10, 13-14, 17-18, 21-22, 25-26, 29-30, 31-34
A mastermix was made for both PCR with
Who was in the lab
Henrike, Jakob, Kristian
Procedure
PCR for 7 fragments:
