Protein Purification: Isolating Fluorescent Protein from Bacteria

In this lab, students use nickel affinity chromatography to isolate fluorescent proteins from E. coli bacteria they have previously transformed with the pKiwi plasmid (or another BABEC plasmid). Nickel affinity chromatography utilizes six histidine (his) amino acids that have been added to the recombinant green fluorescent protein as an artificial “tag.” This 6XHis tag has a high affinity for nickel resin. Since only the recombinant protein will possess this unique tag, the purification process can specifically separate the fluorescent protein from the other proteins in the cell lysate. This lab provides an opportunity for hands on exploration of the applications of genetic engineering, particularly how proteins from GMOs can be isolated for use in research or medicine.

Learning Objectives

  • Isolate purified Green Fluorescent Protein from successfully transformed E. coli bacteria
  • Describe the process of protein purification using nickel-affinity chromatography
  • Create a scientific model that explains how to isolate and utilize a protein product from a genetically modified organism

Prior Knowledge

  • Structure and function of DNA and proteins
  • Relationship between genes and the proteins they code for (DNA → mRNA → protein)
  • Process of bacterial transformation with pKiwi or another plasmid containing a fluorescent protein-coding gene

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Disciplinary Core Ideas:

LS1-A Structure and Function
“…genes are regions in the DNA that contain the instructions that code for the formation of proteins, which carry out most of the work of cells.”

LS3-A Inheritance of Traits
“…genes (expressed) by the cell may be regulated in different ways…”

Science and Engineering Practices:

Developing and Using Models
Constructing Explanations

Students explain how to isolate and utilize a protein product from a genetically modified organism by creating a graphic model.

Crosscutting Concepts:

Systems and System Models

Isolating fluorescent protein from transformed bacteria is a model for protein purification in other applications such as medicine and food production.

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Paradise Valley HS
Phoenix, AZ

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Burton HS
San Francisco, CA

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Lincoln HS
San Francisco, CA

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Burlingame HS
Burlingame, CA

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Marianne Gudmundsson
Glenbrook South HS
Glenview, IL

Well organized and scaffolded; flexible structure and can be used partially or as a whole. This unit can be used as is...superbly written and thought out...incorporates many skills and allows student collaboration and student choice/voice.

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South San Francisco HS
South San Francisco, CA

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Capuchino HS
San Bruno, CA

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