Lab Safety

Safety and security are the prior importance to any work in one’s life. Safety comes first in every small thing that we do as damage plausible can be unpredictable. As it is in life, the same is true in the lab. Lab safety and security are crucial to perform any laboratory activities. Personal safety as well as equipment safety are equally important while working in a lab. To ensure both of these, our team took necessary actions, safety training, and self-awareness about the Institutional Biosafety gdelines. Here are some of the measures we took to ensure Lab safety and security:

Early on in our research endeavors, we participated in a safety training program organized by IISER Tirupati. This program proved invaluable in acquainting us with the various sources of risk and the proper handling and disposal procedures for biohazards, chemicals, and physical hazards. Most of our project work occurred within the undergraduate laboratory at IISER Tirupati.

Throughout our research activities, we remained steadfast in our adherence to both general safety guidelines and, notably, the safety regulations outlined by the biology department. Furthermore, we diligently followed the safety rules and policies set forth by iGEM as they pertained to our research activities in the laboratory. As an additional precaution, we consistently utilized appropriate personal protective equipment, including lab coats, long pants, nitrile gloves, and appropriate footwear.

Waste Disposal

In our laboratory, we diligently manage and segregate the waste generated into distinct categories: Sharp objects, biological solid/plastic waste, biological liquid materials, and other solid waste. Sharps are carefully placed in a puncture-proof container for safe disposal. Biological solid/plastic waste is collected in specially designated red-biohazard bags, subsequently autoclaved for thorough decontamination, and then handed over to the institute's specified agency for incineration. Liquid biological waste is contained in dedicated containers, treated with a 10% bleach solution, and left overnight before being safely discarded. Other solid wastes are gathered and disposed of along with regular waste.

Waste materials that contain hazardous chemicals, such as Ethidium bromide-laden agarose gels and SDS gels, are treated separately. These hazardous waste items are entrusted to the institute's designated agency for incineration. Furthermore, organic solvents and plastic wares contaminated with organic solvents are handled as a separate waste category, distinct from other liquid solvents, to ensure proper disposal.

Organisms, Parts and Activities

During our research activities, we used Escherichia coli DH5Alpha, Escherichia coli BL21, Escherichia coli Nissile 1917 strains which were risk group 1 organisms.

Moreover, All the parts and activities were in the white list.

Responsible Research Practices

To ensure the responsible conduct of our Human Practices, we implemented the following measures:

1. Adherence to Research Ethics: We upheld the principles of social science research ethics, following UNESCO's Code of Conduct for Social Science Research when conducting surveys and in-depth interviews. Additionally, we obtained approval from our institution's ethics committee for our survey activities.
2. Informed Consent: We placed significant emphasis on obtaining written informed consent from all individuals participating in our surveys and in-depth interviews. This consent included:
   • A clear description of the data being collected and its purpose, emphasizing the voluntary nature of participation and the right to withdraw without consequences.
   • A checklist to confirm that participants had the opportunity to ask questions, understood the voluntary nature of their involvement, and agreed to participate in the study or, if applicable, be recorded.
   • We ensured that the informed consent format was in accordance with our institution's guidelines and obtained the necessary approvals.
3. Data Anonymization: We assured survey participants that the collected data would be treated anonymously. For integrated human practices involving experts, we secured written permission for the potential publication of their data. We also considered whether any of the experts were European Union (EU) citizens, enabling us to comply with the General Data Protection Regulation (GDPR).

Bio Safety Kill Switch

We used eLightOn system as the light regulated modules . This system consists of single transcription factors. In the eLightOn system, the synthetic light-switchable repressor LexRO is based on a novel LOV light sensor domain RsLOV.

We constitutively express LexRO. This is LexRO dimerizes and binds to its LexA408 cognate operator sequences and repress the promoter activity. Upon light exposure, the LexRO dimer dissociates causing dissociation from the operator sequences and initiates gene expression. The LexA408 is the cognate sequence incorporated into constitutive promoter CoIE. The toxin and anti-toxin which we will be using is MazF-MazE. MazF is the toxin and MazE is the anti-toxin that is cleaved by either clpA-clpP (or) Lon. This anti-toxin toxin system are involved in regulating cell growth and death under stress conditions. The text explains that MazF is a stable toxin that cleaves mRNAs at ACA sequences, which inhibits protein synthesis and causes cell growth arrest. MazE is a labile antitoxin that forms a complex with MazF and prevents its activity. The method involves a single protein called LexRO that can switch on or off the expression of other genes depending on whether it is exposed to blue light or not.