[예방약학실험8] Luciferase assay

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1. Luciferase assay

Luciferase assay는 NF-κB promoter와 luciferase reporter gene을 포함하는 plasmid를 HEK293 세포에 transfection하여 luciferase 발현을 통해 염증반응 조절에 중요한 유전자 발현을 확인하는 실험입니다. NF-κB는 염증반응 관련 유전자의 신호전달에 관여하는 전사인자로서 염증반응의 지표로 활용됩니다. 실험 결과, PMA는 염증을 유발하고 추출물 A는 항염증 효과가 있음을 확인할 수 있었습니다. GFP로 normalization을 하여 세포 수 차이를 보정하였고, luciferase와 luciferin의 생화학발광반응을 통해 유전자 발현을 정량화할 수 있었습니다. LPS 대신 PMA를 사용한 이유는 HEK293 세포에서 PMA가 NF-κB 등 전사를 활성화하여 염증을 유발하는 것으로 알려져 있기 때문입니다.

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1. Luciferase assay

The luciferase assay is a widely used technique in molecular biology and biochemistry to measure the activity of the luciferase enzyme. Luciferase is an enzyme that catalyzes a bioluminescent reaction, producing light that can be detected and quantified. This assay is commonly used to study gene expression, protein-protein interactions, and various cellular processes. The luciferase assay works by introducing a luciferase reporter gene into cells or organisms, which then expresses the luciferase enzyme. When a substrate (luciferin) is added, the luciferase enzyme catalyzes a reaction that produces light. The amount of light produced is proportional to the activity or expression level of the luciferase enzyme, which can be measured using a luminometer. The luciferase assay is a powerful tool because it is highly sensitive, allowing for the detection of even small changes in gene expression or protein activity. It is also non-invasive, as the light production can be measured without disrupting the cells or organisms being studied. Additionally, the luciferase assay can be used in a wide range of applications, including studying the regulation of gene expression, monitoring cellular signaling pathways, and screening for drug candidates. Overall, the luciferase assay is an essential technique in modern molecular biology and biochemistry, providing researchers with a valuable tool for understanding complex biological processes and developing new therapeutic interventions.