Team:Osaka/Project

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Contents 1 Background 2 Project Details 2.1 System construct 2.2 Detection of DNA damage 2.3 Damage tolerance 3 Results 4 References

Background

In the modern world, radioactivity is a phenomena well understood by science, and harnessed by engineering for nuclear power generation, radiology in medicine, radioisotopic labeling in biological studies and numerous other applications. However, it is an undeniable fact that the ionizing radiation resulting from radioactive decay may cause critical effects on biological creatures, from the smallest bacteria to plants and animals including humans.

On March 11, 2011, the Great East Japan Earthquake struck off the coast of Eastern Japan and triggered a series of equipment failures, nuclear meltdowns, and releases of radioactive materials at the Fukushima 1 Nuclear Power Plant, leading to a nationwide nuclear crisis. While ‘Grays’, ‘Sieverts’, ‘exposure’, ‘equivalent dosage’ and other related terms and units became referred daily in the media, much of the general populace remained ignorant of their meanings and significance. The need for low-cost, portable and easy-to-use dosimeters was apparent as measurements of radiation exposure could only be conducted at dedicated installations spaced far apart and the numbers reported only infrequently.

Hence, this year, iGEM Osaka decided to tackle the building of a biological dosimeter. We attempted to modularly transfer radiation-response and DNA repair genes from the extremophilic bacterium Deinococcus radiodurans to the well-characterized, easily cultivable chassis of Escherichia coli. From there our efforts branched out into (1) evaluating radioresistance conferred by the heterologous genes, and (2) connecting the radiation/DNA damage response system to visible outputs such as color pigment production.

Project Details

System construct

If DNA is damaged greatly by some factors such as UV radiation or chemicals, synthesizes of some damage-related proteins occur quickly. This reaction to DNA damage is SOS response.

RecA is a 38 kilodalton Escherichia coli protein essential for the repair and maintenance of DNA. RecA has multiple activities, all related to DNA repair. In the bacterial SOS response, it has a co-protease function in the autocatalytic cleavage of the LexA repressor and the λ repressor. LexA is expressed constitutively and prevents damage-related proteins to be expressed by binding to SOS box as a repressor. RecA is activated by binding to single-strand DNA caused by damage, and then, conducts activation of LexA protease. Decreasing LexA enable damage-related protein to be expressed.

According to this reaction of bacteria, we employ RecA as a key protein of our system.

Detection of DNA damage

RecA promoter(パーツ名) was employed to detect DNA damage. This promoter induces expression of its downstream genes by DNA damage. Bio-dosimeter must show visible sign as some signal of radioactive substance, therefore we decided to employ pigments as a reporter.

Damage tolerance

Results

References

1. • PprA: a novel protein from Deinococcus radiodurans that stimulates DNA ligation