Alkylating agents comprise a diverse group of chemical compounds. Their sources include industrial chemicals, environmental contaminants, naturally occurring compounds, chemotherapeutic drugs, and experimental carcinogens. In addition to these exogenous sources, there is strong evidence that DNA alkylating species are also produced endogenously. Apart from this, certain alkylating agents such as nitrosamines are also present in potable water and processed food such as cheese. One such specific nitrosamine found is NDMA which is capable of forming lesions in DNA. This suggests that unknowingly we consume these alkylating agents from different sources. But, why should they bother us? These agents are detrimental as they are majorly toxic to bone marrow and gastrointestinal tract. Some of these agents are considered to be potent carcinogens and mutagens. If therapeutically given in high doses, they can lead to sinusoidal obstruction syndrome. Therefore, detection of these alkylating agents becomes imperative.
We propose to create a biosensor, in the form of a bio-sensing solution, that can detect the presence of these alkylating agents. The major reason for us to shift the direction of our project towards the development of a biosensor for detection was that in the recent past we have seen ample publications portraying evidence about whole cell biosensors for detection of analytes like heavy metals, pesticides and other chemicals that alter or influence the biochemical pathway inside the cell. These sensors have the ability to detect concentrations as low as 1micromolar (Mulchandani et al, 2001). Therefore, they are exceedingly sensitive to accurately detect analytes at low concentrations.
Furthermore, we have also proposed a biobrick that can not only detect but also degrade one such alkylating agent, NDMA which is a nitrosamine.
Propitious results in our future experiments would allow us to scale up the production of this biosensor to a commercially viable level and develop it in the form of a dipstick or paper- based kit. Mass production of a microbe like E.coli is cheaper in comparison to the exorbitant scaling up of mammalian cells. Also, these dipsticks or paper-based kits are quite advantageous in lieu of their ease of handling as they do not require any trained personnel for its use.
Our chassis, E.coli- Dh5alpha, strain is a Risk group 1 non-hazardous microorganism and requires Biosafety level 1 laboratory conditions as per the safety guidelines of iGEM. Therefore, addressing the apprehensions of using live cells for detection and subsequent degradation will be relatively uncomplicated.
Priti Mulchandani, Wilfred Chen, Ashok Mulchandani, Joseph Wang, Liang Chen, Amperometric microbial biosensor for direct determination of organophosphate pesticides using recombinant microorganism with surface expressed organophosphorus hydrolase, Biosensors and Bioelectronics,Volume 16, Issues 7–8, 2001, Pages 433-437, ISSN 0956- 5663, https://doi.org/10.1016/S0956-5663(01)00157-9.