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Team:Warsaw/Calendar-Main/23 July 2009
From 2009.igem.org
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Cloning of mitochondrial targeting signal
Sebastian
Task 1:
- PCR with gradient temperature of anniling
Methods:
- Reaction mixture:
1ul pAcGFP1-Mito (5ng) 1ul dNTPs (2mM) 1ul L-primer (5pmol/ul) 1ul R-primer (5pmol/ul) 1ul Pfu buffer 4,5ul ddH20 0,5ul Pfu DNA polymerase
- PCR program:
5min 95C 1min 95C 1min gradient 48-60C 1min 72C go to 2-nd step (30x) 5min 72C
Cloning of the mgtc promoter into the pKSII+ plasmid
Kamil
Tasks:
- Transformant selection
Methods:
- The colonies that remained white after 48h were picked up and transferred to a new petri dish.
- Liquid cultures were established along the way for plasmid purification.
- Both the dish and the liquid cultures were incubated at 37°C overnight.
Assembly of endosomal detection operon
Marcin
Task 1:
- Prepare the bacterial cultures for plasmid isolation
Preparation of bacterial cultures
- Prepare LB medium with kanamycin
- Add 3.5 ml of the medium to the probes
- Add one bacterial colony to each probe
- Breed the bacteria about 7 hours
Construction of K177012 operon1_part2
Ania
Tasks:
- Transformation of the chemocompetent cells with the ligation of BBa_B0032- RBS.3 on the pSB1A2 ampicillin resistant plasmid and - lacI repressor
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