pDsRed-Monomer-Golgi

pDsRed-Monomer-Golgi encodes a fusion protein consisting of DsRed-Monomer, a monomeric mutant derived from the tetrameric Discosoma sp. red fluorescent protein DsRed (1), and a sequence encoding the N-terminal 81 amino acids of human beta 1,4-galactosyltransferase (GT; 2). This region of human beta 1,4-GT contains the membrane-anchoring signal peptide that targets the fusion protein to the trans-medial region of the Golgi apparatus (3–5).

pDsRed-Monomer-Golgi encodes a fusion protein consisting of DsRed-Monomer, a monomeric mutant derived from the tetrameric Discosoma sp. red fluorescent protein DsRed (1), and a sequence encoding the N-terminal 81 amino acids of human beta 1,4-galactosyltransferase (GT; 2). This region of human beta 1,4-GT contains the membrane-anchoring signal peptide that targets the fusion protein to the trans-medial region of the Golgi apparatus (3–5).

SV40 polyadenylation signals downstream of the DsRed-Monomer-Golgi fusion direct proper processing of the 3' end of the mRNA. The vector backbone also contains an SV40 origin for replication in mammalian cells expressing the SV40 T-antigen. A neomycin resistance cassette (Neor ) consisting of the SV40 early promoter, the neomycin/kanamycin resistance gene of Tn5, and polyadenylation signals from the herpes simplex virus thymidine kinase (HSV-TK) gene allow stably transfected eukaryotic cells to be selected using G418 (7). A bacterial promoter upstream of this cassette drives expression of the gene encoding kanamycin resistance in E. coli. The pDsRed-Monomer-Golgi backbone also provides a pUC origin of replication for propagation in E. coli and an f1 origin for single-stranded DNA production.