Figure 1

Description of vector pMJ-LEXSY-rFc. Top: DNA and protein sequence of cloning site and tag region of pMJ-LEXSY-rFc. Bottom: Vectormap of pMJ-LEXSY-rFc. The T7/Tet region (blue) designates the T7 promoter under control of a Tet operator. LMSAP (black) is the secretory leader-peptide of Secreted Acid Phosphatase 1 found in Leishmania Mexicana. The scFv A10 (red) is inserted via Nco I and Not I. The Fc-encoding region of rabbit IgG (turquoise) including the IgG linker (green) is placed just downstream of Not I followed by a Tev restriction site (black), a His-tag (yellow), and a c-Myc-tag (pink) encoding region. The bleomycin resistance gene is fused with a cherry gene, hence designated BleCherry. This combination allows for selection of recombinants with bleomycin and subsequent screening of the best expressing clones by monitoring the cherry fluorescence. The Pci I restriction site is used for linearization and is placed in a telomere region (LTE + RTE), which upon transformation stabilises the linear episome. Finally, the vector holds a tandem T7 transcription terminator (2xT7), a bacterial origin of replication (PBR322 ori), and the Bla ampicillin resistance marker.