2.1 Expression and purification of recombinant
proteins
Triple M2e peptides derived from human (hM2e), swine (sM2e) and avian
(aM2e) IAV were combined with the C-terminus of Cap protein in different
arrangements using Gly-Gly-Gly-Gly linker. (Figure 1a-1c). The cysteine
of M2e were mutated to serine. These sequences were inserted into the
pET28a vector by BamHI and HindIII digestion enzymes, and transformed
into E. coli BL21 (DE3). Then these transformed cells were
induced expression at 20°C for 15 h by isopropyl-β-d-thiogalactoside
(IPTG) (0.2 mM). These recombinant proteins were purified by using
Ni-NTA His·Bind Resin and identified by SDS-PAGE and Western blot. The
concentrations of these purified proteins were determined with a BCA
protein assay kit. The endotoxin concentrations were measured by
ToxinSensor Single Tests Kit.