A Laboratory Exercise For Visible Gel Filtration Chromatography Using Fluorescent Proteins

作者：Zhang Wenqiang; Cao Yibin; Xu Lishan; Gong Jufang; Sun Meihao^*

来源：Biochemistry and Molecular Biology Education, 2015, 43(1): 33-38.

DOI：10.1002/bmb.20833

• gel filtration chromatography
• fluorescent protein
• laboratory exercise

Gel filtration chromatography (GFC) separates molecules according to size and is one of the most widely used methods for protein purification. Here, red fluorescent protein (RFP), green fluorescent protein (GFP), yellow fluorescent protein (YFP), cyan fluorescent protein (CFP), and/or their fusion proteins were prokaryotically expressed, purified, and used in a laboratory exercise to intuitively demonstrate GFC. Different bands, corresponding to RFP, RFP-CFP (RC), YFP-RFP-YFP (YRY), and pyruvate kinase II-GFP (PKG) were well separated on a Superdex 200 column from a 0.5-mL sample. Increasing the sample volume and changing the chromatographic resin to Sephadex G-100 resulted in lower resolution separation. Students enjoyed identifying combinations of colored proteins and found this exercise helpful for understanding the factors that affect GFC resolution.

• 出版日期2015-2
• 单位浙江师范大学