Team:Calgary/InterLab/imposter

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  • Interlab

The InterLab Study

Introduction

The interlab study is an international collaborative lab study where labs around the globe perform identical procedures so that synthetic biology can establish reliable and repeatable measurements and procedures. This year, the interlab study focused on GFP expression in the E. coli strain DH5ɑ using different ribosome binding sites and promoters. GFP was measured via fluorescence readings on a plate reader, and a standard measurement procedure was given out by iGEM that all participating teams followed. This universal protocol is critical, as it enables data to be compared with as many constant variables as possible to ensure that comparisons are accurate.

This study involved two different ribosome binding sites tested with three different promoter regions, along with a positive and negative control, for a total of 8 different trials. Each trial was compared based on its GFP fluorescence readings. The procedure that we followed for this study, which was distributed by iGEM,can be found here

Provided below are the plasmid diagrams of each trial, as well as a brief summary of what each part does. All parts came from the iGEM registry, and can be studied more thoroughly on the iGEM site by clicking on their names below. Also provided are the graphs showing the absorbance and fluorescence readings from the experiment.

Parts & Plasmids

	BBa_E0040: This part is common across all of the transformed plasmids in the 2017 interlab study (minus the negative control), and codes for the production of GFP. GFP, or green fluorescent protein, is a protein native to the Aequeora victoria jellyfish, which glows green when exposed to UV light. GFP is used here as it provides a visual result that can be measured to compare expression of the GFP gene amongst the various plasmids.
	BBa_0010 and BBa_0012: Another couple of parts that are common to all interlab plasmids, BBa_0010 and BBa_0012 are both transcriptional terminators that stop RNA polymerase from transcribing DNA into RNA. Simply put, these parts finish the genetic circuit.
	Chloramphenicol resistance: This part gives resistance to the antibiotic chloramphenicol, allowing bacteria containing this plasmid to be grown on chloramphenicol-containing agar plates.
	BBa_B0032: This part codes for a ribosome binding site, the location where ribosomes bind to begin protein translation. B0032 is considered a weak binding site when compared to B0034, and is only used in the positive control, not in any actual tests.
	BBa_B0034: This part codes for a ribosome binding site, the location where ribosomes bind to begin protein translation. B0034 is one of the two ribosome binding sites tested in this experiment.
	BBa_J364100: This part codes for a ribosome binding site, the location where ribosomes bind to begin protein translation. J364100 is one of the two ribosome binding sites tested in this experiment.
	BBa_J23151: This part is a promoter region, which recruits RNA polymerase to begin transcription of DNA to RNA. J23151 is only used in the positive control, not in any of the actual tests.
	BBa_R0040: This part is a promoter region, which recruits RNA polymerase to begin transcription of DNA to RNA. R0040 is repressed in the presence of tetracycline, and is only used in the negative control, not in any of the actual tests.
	BBa_J23101: This part is a promoter region, which recruits RNA polymerase to begin transcription of DNA to RNA. J23101 is one of the three promoters tested in this experiment.
	BBa_J23106: This part is a promoter region, which recruits RNA polymerase to begin transcription of DNA to RNA. J23106 is one of the three promoters tested in this experiment.
	BBa_J23117: This part is a promoter region, which recruits RNA polymerase to begin transcription of DNA to RNA. J23117 is one of the three promoters tested in this experiment.

Positive Control

Negative Control

Test Device 1

Test Device 2

Test Device 3

Test Device 4

Test Device 5

Test Device 6

Graphs of results

References

Rose, C., Parker, A., Jefferson, B., & Cartmell, E. (2015). The Characterization of Feces and Urine: A Review of the Literature to Inform Advanced Treatment Technology. Critical Reviews In Environmental Science And Technology, 45(17), 1827-1879. http://dx.doi.org/10.1080/10643389.2014.1000761