Safety is always an important consideration in any job and in any product. When it comes to genetically modified organisms, safety is the primary and acute issue. PLASTID PESTICIDES™ project aims to use plastisome-mediated RNAi for efficient control of Spodoptera litura. PLASTID PESTICIDES™ consider a variety of possible safety issues for PLASTID PESTICIDES™ products. This year, PLASTID PESTICIDES™ analyzed the risks that the entire project could encounter. It is also elaborated from two aspects: laboratory safety and gene diffusion safety.
In addition to meeting stringent safety testing requirements, PLASTID PESTICIDES™ team members receive comprehensive training in laboratory techniques that cover a wide range of techniques commonly used in basic experiments. During the training, PLASTID PESTICIDES™ learned:
PLASTID PESTICIDES™ university and college have established strict safety. Regulations that PLASTID PESTICIDES™ follow (see links below):
(https://chemmat.hubu.edu.cn/info/1090/1414.htm)
PLASTID PESTICIDES™ have attached photos showing the training sessions and the implementation of safety protocols.
"The images above depict PLASTID PESTICIDES™ training in strict safety regulations and the implementation of safety protocols, conducted by Professors Shengchun Li and Zhifan Yang, Senior Technician Sheng Yang, and Professor Haimou Zhang."
After official guidance and PLASTID PESTICIDES™ in-house training, PLASTID PESTICIDES™ now have a clear understanding of the difference between a biosafety cabinet and a clean bench. A biosafety cabinet is a device designed to handle potentially infectious materials, and it is equipped with a high-efficiency particulate air (HEPA) filter that filters incoming and expelling air. This design helps protect operators from exposure to particles and aerosols generated by biologics, thus protecting people and the environment. It is important to note that biosafety cabinets are not intended for use in the handling of volatile or flammable chemicals.
PLASTID PESTICIDES™ understand that clean benches are primarily used to create a sterile work environment, but they do not provide protection for the human body. Therefore, PLASTID PESTICIDES™ recognize that only microbial-related operations in a biosafety cabinet can ensure the safety of operators while preventing potential contamination and injury.
The molecular experiments in PLASTID PESTICIDES™ project mainly use safety standard kits and non-toxic reagents as much as possible. PLASTID PESTICIDES™ will comply with all laboratory disciplines and academic standards, and conduct all experiments under guidance and safety precautions, including wearing experimental clothes, gloves and masks throughout the experiment, conducting electrophoresis in designated areas, and washing hands after completing all experiments. In addition, PLASTID PESTICIDES™ will not take any products out of the laboratory. Any activity that may violate the rules and biosafety regulations is prohibited. After the experiment, PLASTID PESTICIDES™ will strictly disinfect the experimental site to avoid any bacteria escaping from the laboratory.
To prevent S. litura escape into the environment:
In addition, PLASTID PESTICIDES™ have mastered the correct disposal methods for different types of waste, and learned how to effectively reduce the risk of contamination and ensure a clean and safe laboratory environment. Through these efforts, PLASTID PESTICIDES™ have not only improved PLASTID PESTICIDES™ safety awareness and operational skills, but also laid a solid foundation for ensuring the smooth progress of scientific research.
PLASTID PESTICIDES™ have detailed waste management procedures that comply with university regulations (see link below):
[University Waste Management Regulations]
(https://xxgk.hubu.edu.cn/info/1244/2002.htm)
- Plants Collection: Carefully collect the GM plant materials, ensuring that all parts including leaves, stems, and roots are included.
- Autoclave bag: Put the plant materials into an autoclave bag.
- Sealing: Securely seal the autoclave bag to prevent any leakage or exposure to the environment during the sterilization process.
- Equipment Setup: Ensure that the autoclave is in good working condition and properly calibrated. An autoclave is a device used for sterilization using saturated steam under pressure.
- Loading: Load the sealed containers into the autoclave chamber. Avoid overloading the autoclave to ensure proper steam circulation and even temperature distribution.
- Temperature and Pressure Settings: Set the autoclave to reach an internal temperature of 121 degrees Celsius (249.8 degrees Fahrenheit) with a corresponding pressure of approximately 15 pounds per square inch (psi).
- Cycle Duration: The sterilization cycle typically lasts for a minimum of 20 minutes. However, the duration may vary based on the volume and type of plant materials being sterilized.
- Monitoring: Continuously monitor the temperature and pressure throughout the sterilization process to ensure that the conditions are maintained within the specified range.
- Venting: After the sterilization cycle is complete, allow the autoclave to cool down naturally. Do not manually vent the autoclave as this could result in uneven cooling and potentially compromise the sterility.
- Unloading: Once the autoclave has cooled down to a safe temperature, carefully unload the containers from the autoclave.
- Disposal: Treat the sterilized plant material as biohazardous waste and dispose of it according to local regulations and guidelines. This might involve incineration or deep burial.
- Documentation: Record the details of the sterilization process, including the date, time, temperature, pressure, and duration. This documentation is important for regulatory compliance and traceability.
By following these steps, PLASTID PESTICIDES™ can effectively sterilize the genetically modified plants at 121 degrees Celsius to ensure that they are completely killed and will not inadvertently enter the environment. This process is crucial for maintaining biosafety and preventing unintended ecological impacts.
In order to effectively manage potential safety risks, PLASTID PESTICIDES™ have taken a variety of measures, including writing detailed incident reports, actively participating in safety seminars sponsored by iGEM, and proactively engaging with industry experts. On top of this, PLASTID PESTICIDES™ have also worked together to develop safe practices in the laboratory to minimize potential threats to laboratory staff and the surrounding environment. PLASTID PESTICIDES™ project to study Spodoptera liturae has passed a safety audit.
https://teams.igem.org/5044/safety
Plastids (chloroplasts) are inherited matrilineally in most crops. Matrilineal inheritance excludes plastid genes and transgenes from pollen transmission. Plastids exhibit maternal inheritance, meaning they are not present in pollen, largely excluding plastid-localized transgenes from pollen transmission, greatly reducing the risk of unwanted transgenes being transferred from genetically modified (GM) plant fields to adjacent fields of non-GM plants. This alleviates environmental safety concerns. Therefore, plastid transformation is considered to be an excellent tool to ensure transgenic containment and improve the biological safety of transgenic plants.
The application of the PM-RNAi method in insect management has significant environmental advantages, especially in the protection of pollinators and non-target insects. Since pollen sperm cells exclude plastids in angiosperms, this means that the risk of GMOs spread through pollen is naturally controlled. As a result, pollinators and pollen-feeding insects are not exposed to amiRNA produced by plastids, reducing the potential impact on these non-target organisms.
Artificial miRNAs (amiRNAs) are computationally generated sequences with random arrangements that resemble naturally occurring miRNAs. They are designed to reroute endogenous miRNA biogenesis and silencing mechanisms in order to generate highly specific and efficient transient or stable loss-of-function transformants for target genes.
Gene silencing of artificial microRNAs is one of the most critical methods for developing desired horticultural plants. amiRNAs are highly specific, approximately 21 nt in length, and are designed to inhibit post-transcriptional gene expression. amiRNA-based strategies are considered more effective than siRNAs because of their high specificity and minimal off-target effects. In contrast, artificial microRNAs have high sequence specificity and can only recognize less than 5 mismatched target sequences. In this way, it is not easy to target non-target genes without off-target.
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Genetically modified organisms are living organisms whose genetic material has been altered using genetic engineering techniques. These modifications can include the insertion, deletion, or alteration of genes to produce desired traits, such as resistance to pests, improved nutritional value, or increased yield.
Plastisomes are organelles within plastids, such as chloroplasts, which contain the machinery necessary for protein synthesis. They play a crucial role in the expression of plastid-encoded genes and are involved in the production of proteins required for photosynthesis and other plastid functions.
Laboratory hazards are potential sources of harm in a laboratory setting. These can include chemical, biological, physical, and ergonomic hazards. Proper identification and management of these hazards are essential to maintaining a safe working environment.
A biosafety cabinet is an enclosed, ventilated workspace designed to provide personnel, environmental, and product protection. It uses HEPA filters to purify the air and is used when handling potentially infectious agents or hazardous substances.
A clean bench is a laminar flow work station that provides a sterile, particle-free work area. Unlike a biosafety cabinet, it does not offer protection to the user; instead, it protects the work from contamination.
An autoclave is a device that uses pressurized steam to sterilize equipment and supplies. It is commonly used in laboratories and healthcare settings to eliminate microorganisms, including bacteria, viruses, fungi, and spores.
Matrilineal inheritance is a pattern of heredity in which certain genetic traits are passed down from mother to offspring. In plants, this often applies to plastids, which are inherited through the female line, excluding them from pollen transmission and reducing the spread of transgenes.
Artificial microRNAs are synthetic RNA sequences designed to mimic natural microRNAs. They are used to silence specific target genes by integrating into the endogenous miRNA pathway, leading to the degradation of the target mRNA or inhibition of its translation.
Gene silencing is the regulation of gene expression in a cell to prevent the activity of a specific gene. It can occur naturally through mechanisms like RNA interference, or it can be induced artificially, for example, by introducing amiRNAs.
Transgene containment refers to the strategies and measures taken to prevent the unintentional spread of genetically modified genes from GM organisms to non-GM organisms. This is important for preventing unintended ecological impacts and maintaining the integrity of non-GM crops.
Biological safety involves the practices and procedures implemented to protect individuals and the environment from exposure to potentially harmful biological agents. This includes the use of proper containment, handling, and disposal methods for biological materials.