Figure 5
From: Development of novel detection system for sweet potato leaf curl virus using recombinant scFv
Expression test and avidity effects of bivalent scFv. (A) Plasmid vector map for bivalent scFv. This construct was designed based on a MBP fusion expression plasmid, pDEST-periHisMBP64. The existing F7 gene (shown in yellow) and the E. coli codon-optimized F7 gene (shown in green) were ligated with a glycine-serine linker (shown in blue). The AflII and HindIII restriction enzyme sites that do not digest the backbone were added, so that the linker could be replaced. (B) Expression test of bivalent F7 scFv protein. The molecular weight of the expressed fusion proteins was 103.34 kDa in total and the molecular weight of each subunit was as follows: signal peptide (N-terminal signal peptide of MBP; 4.6 kDa) and MBP and polyhistidine tag (45.3 kDa). Lane 0 shows non-induction, lanes 1–3 show induction with IPTG (0.1, 1, and 2 mM). The induction was carried out at OD600 = 0.6, and overexpressed at 26 °C for 16 h. (C) Bivalent scFv (●) and monovalent scFv (■) were bonded with serial dilution in homogenized sweet potato leaf curl virus-infected leaf samples in coated ELISA wells. The 3D8 scFv (▲) was used as a negative control. Data are presented as means ± SEMs.
