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Team:Bielefeld-Germany/Labjournal/week13
From 2012.igem.org
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* '''Team Modeling''' and '''Team Sponsoring''': meeting Mr. ??? from the clarification plant of Schloß Holte and finding a new cooperation partner. He want to give us the information we need to equate our model and design our cleaner. On top he wants to ask if the clarification plant can sponsor our project. | * '''Team Modeling''' and '''Team Sponsoring''': meeting Mr. ??? from the clarification plant of Schloß Holte and finding a new cooperation partner. He want to give us the information we need to equate our model and design our cleaner. On top he wants to ask if the clarification plant can sponsor our project. | ||
| + | * '''Team Site Directed Mutagenesis:''' Went on with the PCR-products and transformed them into XL1 Blue | ||
===Friday July 27th=== | ===Friday July 27th=== | ||
Revision as of 20:44, 19 August 2012
Contents |
Labjournal
Week1 Week2 Week3 Week4 Week5 Week6 Week7 Week8 Week9 Week10 Week11 Week12 Week13 Week14 Week15 Week16 Week17 Week18
Week 13 (07/23 - 07/29/12)
Monday July 23rd
- Some of our team members were at CAS conference in Munich from 07/23 - 07/25/12.
- Successful PCRs of laccase gene lac5 from Trametes versicolor with plasmid DNA from Uni Greifswald as template.
- Team Site Directed Mutagenesis: Transfered 4 colonys per petri dish to an new one for plasmidisolation
Tuesday July 24th
- Team Activity Tests: Today was the moment of truth. We received the first recombinant laccases from the cultivation team. We started with testing the supernatant from the cultivated cells to check whether they secrete laccase. We added 0,1 mM ABTS in each well that was filled with supernatant from Escherichia coli, Bacillus pumilus, Bacillus halodurans C-125, Xanthomonas campestris pv. campestris B100 and E. coli KRX (negative control - cells without plasmid) and measured as usual. There was no change in the OD so that we assume that no secretion takes place. Afterwards we tested potential laccase activity by filling each well with laccase and buffer (check protocols to see which buffer we used) and added 0,1 mM ABTS. Unfortunately no activity was seen. We will try again soon.
- Team Site Directed Mutagenesis: Plasmidisolation of B.pumilus 2883(I-IV); X.campestris 2247(I-IV); T.thermophilus 2796(I-IV) and E.coli 2307 (III).
Wednesday July 25th
- Team Site Directed Mutagenesis: Testrestriktion of the Plasmids revealed that B.pumilus III;IV; E.coli III;T.thermophilus.I;II seem to be mutated corrcetly but X.campestris is untouched (no Band at 3636).
- SDM-PCR with B.pumilus III(III)+IV(IV) as template using the SDM-primermix to mutate bp 2317
- SDM-PCR with X.campestris with the 2247-primermixand another one with the primermix to mutate 3633
- Prepared Testrestrikion-positives for Sequencing
Thursday July 26th
- Team Modeling and Team Sponsoring: meeting Mr. ??? from the clarification plant of Schloß Holte and finding a new cooperation partner. He want to give us the information we need to equate our model and design our cleaner. On top he wants to ask if the clarification plant can sponsor our project.
- Team Site Directed Mutagenesis: Went on with the PCR-products and transformed them into XL1 Blue
Friday July 27th
Saturday July 28th
- Team Activity Tests: See, we are back already! Today we received different samples from Escherichia coli, Bacillus pumilus, Bacillus halodurans C-125, Thermus thermophilus, Xanthomonas campestris pv. campestris B100 and E. coli KRX (negative control - cells without plasmid). The cells were all cultivated at 30°C and supplied with copper. We tested each one of them but none of them showed laccase activity.
