Schematic representation of bicistronic retroviral vectors. The ABCA1 cDNA was inserted behind a spleen focus-forming virus (SFFV) retroviral LTR following the splice donor (SD) site. The neomycin resistance (neo) cDNA was inserted after a splice acceptor (SA) site derived from the genome of the Moloney murine leukemia virus. Hemagglutinin (HA) epitope was introduced into the first extracellular loop of the N-terminal transmembrane domain (TMD) of ABCA1 (between residues 207 and 208). WT: wild type untagged ABCA1 construct; HA-WT: wild type HA-tagged ABCA1 construct; HA-MK: a variant with a methionine-lysine substitution (K939 M) in the Walker A motif of the N-terminal nucleotide-binding domain (NBD1); HA-KM: methionine-lysine substitution (K1952 M) in the C-terminal nucleotide-binding domain (NBD2); HA-MM: methionine-lysine substitution in both Walker A motifs (K939 M/K1952 M). A vector containing only the neo cDNA after the SA site was used as a negative control (vector).
Kasza et al. BMC Cell Biology 2009 10:93 doi:10.1186/1471-2121-10-93