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Yeast two-hybrid assay with fluorescence reporter
Korean J. Microbiol 2019;55(3):199-205
Published online September 30, 2019
© 2019 The Microbiological Society of Korea.

Seong Kyun Park, Su Ryeon Seo*, and Byung Joon Hwang*

Department of Molecular Bioscience, College of Biomedical Science, Kangwon National University, Chuncheon 24341, Republic of Korea
Correspondence to: (B.J. Hwang) E-mail: bjhwang@kangwon.ac.kr; Tel.: +82-33-250-8543; Fax: +82-33-259-5641 /
(S.R. Seo) E-mail: suryeonseo@kangwon.ac.kr; Tel.: +82-33-250-8541; Fax: +82-33-259-5641
Received August 2, 2019; Revised August 27, 2019; Accepted August 27, 2019.
Abstract
Yeast two-hybrid (Y2H) technique has been used to study protein-protein interactions, but its application particularly to a large-scale analysis of protein interaction networks, is limited by the fact that the technique is labor-intensive, based on scoring colonies on plate. Here, we develop a new reporter for the measurement of the protein-protein interactions by flow cytometry. The yeast harboring interacting proteins can also be enriched by fluorescence-activated cell sorting (FACS) or magnetic-activated cell sorting (MACS). When two interacting proteins are present in the same yeast cell, a reporter protein containing 10 tandem repeats of c-myc epitope becomes localized on the surface of the cell wall, without affecting cell growth. We successful measured the surface display of c-myc epitope upon interacting p53 with SV40 T antigen by flow cytometry. Thus, the newly developed Y2H assay based on the display of c-myc repeat on yeast cell wall could be used to the simultaneous analysis of multiple protein-protein interactions without laborious counting colonies on plate.
Keywords : flow cytometry, fluorescence reporter, protein-protein interaction, yeast two-hybrid


September 2019, 55 (3)
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