Figure 2

Cloning sites and fusion products. (A) pGBKCg generates a C-terminal fusion of the Gal4 DBD separated by a 13 amino acid linker from the N-terminally fused ORF (here: VZV ORF 19). A MYC tag is embedded in a C-terminal 37 amino acid tail. (B) pGADCg generates a C-terminal fusion of the Gal4 AD. The cloned ORF (here: VZV ORF 26) is preceded by a 33 amino acid sequence that contains the nuclear localization signal (NLS). The NLS of pGBKC is part of the DBD. A 14 amino acid linker separates the ORF from the Gal4 AD. The HA tag of pGADCg has been shortened to seven amino acids so that it may not be recognized by anti-HA antibodies. During the LR reaction the Gateway cassette in both vectors will be replaced by the sequence of interest plus additional nucleotides from the entry clone (i.e. the underlined sequence will be replaced). For further technical details see the Gateway manuals at http://www.invitrogen.com. pGBKCg and pGADCg have been deposited with and are available from Addgene http://www.addgene.com. Their sequences have been deposited with GenBank under accession numbers FJ696409 (pGBKCg) and FJ696408 (pGADCg). pGBKT7g and pGADT7g are available from the authors.