The information provided below is for popular science and theoretical discussion only and does not constitute any guidance for experimental operation or application. Genetic modification involves complex biological, molecular engineering, ecological ethics and legal supervision issues. Relevant research can only be carried out under strict laboratory safety measures, sufficient risk assessment and permission from relevant departments. The following content outlines some basic principles and ideas about using genetic engineering to achieve bioluminescence in published literature. 1. Principles of bioluminescence a. The luminescence system is usually based on two main components: an enzyme that can catalyze the reaction (for example, luciferase or luminiferase) and its corresponding substrate (usually called luciferin or luciferin substrate). In the reaction, the enzyme reacts with the substrate and releases energy in the form of light under the action of oxygen and other cofactors. b. Autonomous bioluminescence (self-sufficiency) requires not only the introduction of luminescent proteins, but also the synthesis pathway of the corresponding substrate; for example, the lux gene cluster of bacteria contains the entire process from substrate synthesis to enzyme reaction. 2. Choose a suitable luminescent system a. Systems from fireflies, marine organisms (such as the green fluorescent protein GFP of the jellyfish, although GFP itself does not emit light directly, but is used as a fluorescent tag), and luminescent bacteria (such as Vibrio fischeri, etc.) are all candidates. b. In recent years, some studies have tried to introduce the entire bacterial luxCDABE gene cluster into eukaryotic organisms to enable them to produce light autonomously, but they face problems such as energy consumption and substrate supply. 3. Genetic engineering strategy a. Gene construction: • When designing the construction, it is necessary to synthesize or extract the required luminescent gene, and consider codon optimization of the gene sequence so that it can be well expressed in the target plant and animal. • If you want to achieve self-sufficiency, you need to integrate genes related to substrate synthesis, not just luminescent enzymes. b. Selection of regulatory elements: • Select appropriate promoters and regulatory sequences to ensure expression in the target cells at specific times and tissues. For example, photosynthetic cell-specific promoters can be used in plants, and tissue-specific or inducible promoters can be used in animals. • The use of a controllable expression system can regulate the luminescence intensity when needed, reducing the burden on the host cell. c. Gene introduction methods: • For plants: Common methods include Agrobacterium-mediated transformation, gene gun or CRISPR-related technology. • For animals: Gene knock-in can be achieved by using viral vectors (such as lentivirus or adeno-associated virus), electroporation or direct use of CRISPR-Cas9 technology. d. Fusion strategy: • To ensure that the luminescence system is coordinated with the host metabolic environment, further engineering may be required for correct protein folding and cellular localization to ensure sufficient contact and stable expression between the enzyme and the substrate. 4. Experimental optimization and challenges a. Expression efficiency and energy load: The luminescence process requires energy consumption. Excessive expression may bring metabolic burden to animal and plant cells, and the expression level needs to be carefully regulated. b. Substrate synthesis and stability: If the target cell itself does not produce suitable luciferin, it is necessary to introduce a complete substrate synthesis pathway at the same time; the synthesis reaction may be affected by the metabolic state of the cell. c. Ecological and ethical risks: The release of transgenic plants and animals into the natural environment may bring unforeseen ecological effects. Therefore, most studies are limited to strictly controlled laboratory environments and must comply with relevant national and international regulations. 5. Existing research cases a. Some studies have successfully made bacteria and yeast self-luminous under laboratory conditions; recently, there have been reports of introducing luminescent gene constructs into plants to achieve simple continuous luminescence. b. In animals, transgenic mice or fish have achieved in vivo luminescence by introducing luciferase genes, but this is usually limited to specific research fields (such as labeling cells, observing transgenic expression, etc.). In summary, to transform animals and plants to make them glow in the dark, it is usually necessary to: 1. Select a suitable luminescent gene system (such as the lux system of fireflies or bacteria); 2. Optimize the sequence of the selected gene, construct a suitable expression vector, and combine it with a suitable promoter and regulatory elements; 3. Use appropriate transformation methods to introduce the constructed gene into the target biological cells; 4. Conduct strict laboratory testing and environmental assessment to ensure that biosafety and ecological risks are minimized. Finally, it is emphasized again: genetic modification involves safety, ethical and legal issues. Any related experiments must be carried out under the premise of professional institutions, strict supervision and full review. This article is limited to theoretical discussion. Any further research and practice need to be carried out under the guidance of professionals and in an environment that complies with legal regulations.