Development of a Fluorescent Protein-based Laboratory Course for Life Science Students

The flow of genetic information from DNA to proteins is fundamental and crucial for students in the life sciences. To achieve a deeper understanding, practical laboratory courses are desirable, where students can extract and purify both DNA and its corresponding protein products from cells to investigate their properties. However, time constraints in student laboratories limit the scope of individual topics. Therefore, we sought to develop a comprehensive curriculum that covers these related fields within a short timeframe. As a result, we developed an experimental system by introducing the genes for four colors of fluorescent proteins—green, red, cyan, and orange (GFP, RFP, CFP, and OFP, respectively)—into Escherichia coli, allowing each protein to be purified from a small volume of culture. The constructed plasmids feature a high-copy replication origin and an ampicillin resistance gene, enabling the high-level expression of each fluorescent protein fused with a polyhistidine tag. E. coli expressing each fluorescent protein exhibit distinct, easily observable colors, and the expressed proteins can be readily purified using a Ni-affinity column. This article is an English translation of the paper originally published in Japanese in the Bulletin of Hiroshima Institute of Technology. Education, Vol. 19, pp. 61-69 (2020). Available at: https://it-hiroshima.repo.nii.ac.jp/records/453The annotated plasmid sequences (pUC-GFP-H6, pUC-RFP-H6, pUC-CFP-H6, and pUC-OFP-H6) are provided as GenBank format files (.gb).