Team:Goettingen/week18-2

Revision as of 11:12, 22 September 2012 by Rosin (Talk | contribs)


Home Homepage News Team Members Focus Groups Seminars Work Impressions Project Our Project Results Discussion/Outlook Lab Work Flow Notebook Overview Materials Methods Computational Data Bioinformatical Tools iGEM What is iGEM? Parts Submitted Attributions Safety > Göttingen City University of Göttingen Max-Planck-Institute S1-Demo Lab Human Practice Our Human Practice Projects Public and Media Survey Synthetic Biology Day Panel Discussion Flash Coli Sponsoring Our Sponsors Interested in...?
Deutsch / English

#2 Speed Improvement - 18th week

V08_27_1 Miniprep of the tar-promoter constructs

Experiment:
The Minipreps were conducted using the PeqGOLD MiniPrep Kit (Peqlab) according to the user manual.

V08_27_2 Chemical retransformation of the tar-promoter constructs into E. coli BL21

Experiment:
The retransformation was performed as described in the standard protocol.
Observations & Results:
The retransformation was successful since numerous colonies could be obtained except for the empty negative control.

V08_27_3 Chemical transformation of different motility increasing constructs into E. coli MG1655

Observations & Results:
The transformation was not successful. The next morning only one plate (the positive control) featured any visible colonies. However, after several additional hours of incubation also some very small colonies appeared at some other plates. Nevertheless, the efficiency was very low. We suggest that this is normal since MG1655 is no cloning strain and might have thus a much lower transformation efficiency. This idea was supported by our inquiries.

V08_27_4 Preparation of over night cultures

Experiment:
In oder to perform a Real-Time Quantitative Reverse Transcription PCR, over night cultures of the tar-promoter constructs in BL21 were prepared.

Preparative double digestion of the flhDC-promoter constructs and pSB1C3

Experiment:
The flhDC-promoter constructs as well as the vector pSB1C3 still including tar were digested with EcoRI and PstI as described in the protocol. Afterwards, the fragments of the desired size were cut out and frozen in order to extract the DNA from the gel slices the following day.
Observations & Results:
The gel featured clearly visible bands of the expected length and thus the digestion was successful.

V08_19_1 Purification of the the flhDC-promoter constructs and pSB1C3

Experiment:
The restriction product was purified using peqGOLD Gel Extraction Kit (Peqlab) according to the manual.

V08_29_2 Insertion of the flhDC-promoter constructs into pSB1C3

Experiment:
The ligation of the digested flhDC-promoter constructs with pSB1C3 was conducted as described in the protocol.

Miniprep of the flhDC-promoter constructs in pUC18

Experiment:
In order to obtain further plasmid material Minipreps were conducted using the PeqGOLD MiniPrep Kit (Peqlab) according to the user manual.