Team:Imperial College London/Wetlab/Results

=Wet Lab Results= There's a brief description of each wet lab results and you can click through to find more detail on them. For everyday experiments and minor results, please refer to our Notebook.

OD calibration
A calibration curve is generated that relates the OD600 readings with the actual colony forming units when plated on agar plates.

IPTG Growth
The cells are grown in varying amounts of IPTG to test if IPTG has any toxic effect on the cells.

PcstA activity with different media
The activity of the PcstA promoter is monitered by measuring GFP fluorescence after overnight growth in different types of media.

Thermoinducible promoter system
The thermoinducible system with GFP is tested and shown that there is activation of the promoter when the temperature is raised to 42°C.

Restriction Enzyme activity
An experiment investigating the effects of methylation on the amount of cleavage by varying concentrations of restriction enzymes.

=Others=

=Calibration=

OD calibration
OD calibration

23rd September
 * To make a calibration curve linking our O.D. 600 to colony forming units.

2nd October

=Module 1=

=Module 2=

How cell lysis varies with pH (using GFP as marker). Chemically induced encapsulation 18-09
=Module 3=

Restriction Enzyme testing in Dam+ve and Dam-ve Strains
An experiment investigating the effects of methylation on the amount of cleavage by varying concentrations of restriction enzymes.

=Temporal Control=

Cell growth for different IPTG concentrations
Testing if IPTG toxicity affects cell growth, and consequently, production of protein of interest. Rationale:  IPTG has been shown in the past to have toxic effects on cell growth if administered in large quantities (literature review IPTG). However, production of the protein of interest will be induced with IPTG and from the Modelling results, output yield of protein of interest will increase with concentration of IPTG, provided that the levels are non-toxic. The results we present here are a study on how IPTG exerts its effects on cellular growth rate, in order to determine if the concentrations we are using have negative effects on the cultures, and consequently, how this will affect output levels of drug of interest.

Diauxic growth
8th September 10th September

Growth in the presence of Glucose only
Testing the effects of glucose concentration on cell growth: This experiment will be linked to diauxic growth. Rationale: A cell culture in the presence of glucose grows exponentially. Once it uses up its resource, it stops growing and the population density saturates. Raw data 06/10 [[Media:II09_rawdata.xls]]

Growth in the presence of Glucose + 2ndary carbon sources
Testing the effects of different secondary carbon sources on cell growth with a fixed initial glucose concentration. Rationale: A cell culture in the presence of glucose grows exponentially. Once it uses up its resource, it stops growing and the population density saturates. However, in the presence of a secondary carbon source, the cells enter a second exponential phase where they grow and saturate once again when the secondary source has been used up. NOTE: Same raw data file as above!

Growth in the presence of Glucose + Xylose
Testing the effects of different xylose concentrations on cell growth, for a fixed initial glucose concentration: This experiment will be linked to diauxic growth. Rationale: As above

Review on CRP promoter results
Prelim test-Growth on glucose and xylose and effect on CRP promoter 07-10 A simple test of all xylose and all glucose to test if GFP is generated. Found out that GFP levels are about the same and quite low for both xylose and glucose (ie both do not express)

Growth on different media and effect on CRP promoter 10-10
Supposing that Secondary carbon sources repress the CRP promoter, we proceeded to test which media gives the best CRP output. It appears to be 10% Casamino acid. [[Media:II09_CRP responsive 10-10.xls| download raw data]]

Prelim test on GFP and RFP production from construct CRP-GFP-Lac-RFP
CRP promoter appears to be constitutively on, whether Glucose or Xylose is present. Most likely because 0.05% glucose is unable to repress CRP promoter.

CRP promoter and varying glucose 14-10
[[Media:crpglc14-10.xls| Download raw data here]]

Harvard+GFP results 6-10
Testing the effects of temperature on fluorescence output. Rationale: Genome deletion is repressed at low temperatures, and triggered when the temperature rises. Here, we are testing the p-lambda promoter by attaching GFP to it, in order to show how the fluorescence output varies when the temperature rises. [[Media:HVD_raw.xls |raw data]] 6th October 23rd September