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Team:Osaka/Tests
From 2012.igem.org
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=== SOS promoter assay === | === SOS promoter assay === | ||
| - | + | <p>We assayed the promoter of the SOS gene RecA ([http://partsregistry.org/wiki/index.php?title=Part:BBa_J22106 J22106]). | |
To measure the DNA damage detection, we used antibacterial agents as a source of DNA damage. To quantitatively and accurately evaluate the promoter activity, dual luciferase assay method was employed. | To measure the DNA damage detection, we used antibacterial agents as a source of DNA damage. To quantitatively and accurately evaluate the promoter activity, dual luciferase assay method was employed. | ||
Transformed ''E. coli'' was exposed to antibacterial agents and then incubated for 2 hours. For details check the [https://2012.igem.org/Team:Osaka/Protocols Protocols page].</p> | Transformed ''E. coli'' was exposed to antibacterial agents and then incubated for 2 hours. For details check the [https://2012.igem.org/Team:Osaka/Protocols Protocols page].</p> | ||
Revision as of 02:40, 11 September 2012
Tests
Damage tolerance assay
To measure the DNA damage tolerance conferred by each part, we used antibacterial agents as a source of DNA damage and then assayed the survival rates. Transformed E. coli cells were plated on agar plates at different dilutions, air dried, and then exposed to different doses of UV radiation. Plates were wrapped with aluminum foil and incubated in the dark. Colony-forming units were scored after 16h incubation at 37°C. For detailed protocols, refer to the Protocols page.
The tolerance parts tested were as follows:
Parts containing one gene each
- CDS: PprI, PprA, PprM or RecA
Parts containing two genes
- CDS1+2: PprI+RecA, PprA+RecA, PprM+RecA, PprI+PprA, PprI+PprM, PprA+PprM
Discussion
Single-gene parts
Two-gene combinations
Conclusion
SOS promoter assay
We assayed the promoter of the SOS gene RecA ([http://partsregistry.org/wiki/index.php?title=Part:BBa_J22106 J22106]). To measure the DNA damage detection, we used antibacterial agents as a source of DNA damage. To quantitatively and accurately evaluate the promoter activity, dual luciferase assay method was employed. Transformed E. coli was exposed to antibacterial agents and then incubated for 2 hours. For details check the Protocols page.
