Interlab study

Our team has contributed to the Interlab study not only by measuring some of the proposed constructions but also by sharing the results of similar constructions built for our own project. You can find our results with the Interlab construction BBa_I20260 below, and also brief explanations and links to the detailed results of our own -supplementary- constructions. You can see a more detailed explanation of the protocols we used in our Interlab Study Worksheet!

Fluorescence emission by Biobrick part BBa_I20260

We used the famous E. coli strain DH5α as a host for the expression of this construction. Briefly, the strain carrying the Biobrick part in the pSB3K3 vector was striked on LB medium supplemented with kanamycin. A liquid culture was set up in the same medium and grew in agitation (250 rpm) at 37°C until OD600 reached a value around 0,2. The fluorescence of this culture was measured and the results are shown in Figure 1. We used the wild-type DH5α E. coli strain as a control, and performed all the experiments with five independent replica.

Figure 1. Fluorescence emission of DH5α E. coli strain containing Biobrick part BBa_I20260 vs the control non-transformed strain. The average and the standard deviation of five independent biological replicates is shown. Fluorescence intensity (FI) was normalized by the optical density of the cultures measured at 600 nm (OD600).

Other constructions tested

In this project, we have built 10 different constructions composed of a reporter protein under the control of a constitutive or inducible promoter. All the individual parts our construction were made of were taken from the Registry of Standard Biological Parts. We managed to fully experimentally test 6 out of the 10 constructions during the summer. For each one of them, we have analyzed their output in different E. coli strains, which were grown under a range of environmental conditions. In addition, some constructions were tested in combination, this is, co-expressed in the same E. coli cell:

Biobrick part 1. Constitutive promoter J23104 from Anderson's collection + GFP (ZsGreen1). This construction was tested in 6 different E. coli strains. In one of these strains, its output was measured in a range of temperatures, pH conditions, salt concentrations, and UV-irradiation. Bb1 was also tested in cotransformation with Bb2.

Biobrick part 2. Constitutive promoter J23104 from Anderson's collection + RFP (AsRed2). This construction was tested in 6 different E. coli strains. In two of these strains, its output was measured under vacuum conditions and UV-irradiation. Bb2 was also tested in cotransformation with Bb1 and Bb3.

Biobrick part 3. Constitutive promoter J23110 from Anderson's collection + GFP (ZsGreen1). This construction was tested in 6 different E. coli strains. In one of these strains, its output was measured in a range of temperatures, pH conditions, salt concentrations, and a href="https://2014.igem.org/Team:Valencia_Biocampus/Results#[edit]UVRadiation(2)" target="_blank">UV-irradiation. Bb3 was also tested in cotransformation with Bb2.

Biobrick part 4. Temperature-inducible GroE promoter + lacZ. The output of this construction was tested in 6 different E. coli strains.

Biobrick part 5. Doxycyclin-inducible TetR promoter + lacZ. The output of this construction was tested in 6 different E. coli strains.

Biobrick part 6. IPTG-inducible tac promoter + lacZ. The output of this construction was tested in 6 different E. coli strains.