Author: van der Schaar, H. M.; Melia, C. E.; van Bruggen, J. A. C.; Strating, J. R. P. M.; van Geenen, M. E. D.; Koster, A. J.; Bárcena, M.; van Kuppeveld, F. J. M.
Title: Illuminating the Sites of Enterovirus Replication in Living Cells by Using a Split-GFP-Tagged Viral Protein Document date: 2016_7_6
ID: 1aptufp6_9
Snippet: Before introducing GFP(S11) into the viral genome, we first tested whether inserting the tag into 3A at this position would generate GFP fluorescence upon coexpression with GFP(S1-10). Staining with an anti-3A antibody demonstrated that the expression pattern of 3A(S11aa2) resembled that of untagged 3A [i.e., lacking GFP(S11)] in HeLa cells (see Fig. S1A in the supplemental material) and in BGM (buffalo green monkey) cells (data not shown), indic.....
Document: Before introducing GFP(S11) into the viral genome, we first tested whether inserting the tag into 3A at this position would generate GFP fluorescence upon coexpression with GFP(S1-10). Staining with an anti-3A antibody demonstrated that the expression pattern of 3A(S11aa2) resembled that of untagged 3A [i.e., lacking GFP(S11)] in HeLa cells (see Fig. S1A in the supplemental material) and in BGM (buffalo green monkey) cells (data not shown), indicating that the GFP(S11) tag does not alter the localization of 3A. Coexpression of GFP(S1-10) with 3A(S11aa2), but not with untagged 3A, resulted in a GFP signal that greatly overlapped with the 3A pattern generated with the anti-3A antibody (see Fig. S1B , HeLa cells not shown). Furthermore, 3A(S11aa2) was not GFP fluorescent in the absence of GFP(S1-10) (see Fig. S1A ). Thus, GFP(S1-10) can assemble with 3A(S11aa2) to yield GFP fluorescence at the sites where 3A is localized in the cell.
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