K. Aghaiypour - Department of Genomics and Genetic Engineering, Razi Vaccine and Serum Research Institute, Agricultural Research, Education, and Extension Organization, Karaj, Iran
R. Teymourpour - Department of Genomics and Genetic Engineering, Razi Vaccine and Serum Research Institute, Agricultural Research, Education, and Extension Organization, Karaj, Iran

Fragment C is the C-terminal domain of the heavy chain of tetanus toxin that can promote the immune response against the lethal dose of this toxin. Therefore, this portion can be considered as a candidate vaccine against tetanus infection, which occurs by Clostridium tetani. The present study aimed to compare the expression of tetanus toxin fragment C in Escherichia coli  BL۲۱ (DE۳) pLysS cells having a high tolerance to toxins between two different expression vectors, namely pET۲۲b and pET۲۸a, using the sodium dodecyl sulfate polyacrylamide gel electrophoresis and western blot analyses. After DNA extraction from Harvard CN۴۹۲۰۵ strain of C. tetani, the gene of interest was amplified using polymerase chain reaction, and then sequenced and cloned into the expression vectors of pET۲۲b and pET۲۸a, transformed into competent BL۲۱ (DE۳) pLysS cells, and finally expressed using an optimized protocol. The cells were induced with isopropyl β-D-۱-thiogalactopyranoside (IPTG) at four different incubation temperatures (i.e., ۳۷, ۳۳, ۳۰, and ۲۵ °C) and three different incubation times (i.e., ۱, ۲, and ۳ h). Although the SDS-PAGE and western blot analyses confirmed the expression of the recombinant fragment C (r-fragment C) ligated into both of the expression vectors, pET۲۸a showed a higher r-fragment C expression level than the other vector (۳۸.۶۶ mg/L versus ۳۲.۳۳ mg/L, P<۰.۰۵). An optimal expression condition was acquired ۳ h after ۱ mM IPTG induction at ۲۵ °C. The results demonstrated that E. coli BL۲۱ (DE۳) pLysS as an expression host in combination with pET-۲۸a as an expression vector was a more compatible expression system to express the fragment C of tetanus toxin, compared to E. coli BL۲۱ (DE۳) pLysS/pET-۲۲b expression system. Overall, these results may represent an opportunity to improve the expression system for the production of tetanus toxin vaccine using recombinant protein strategy.

Clostridium tetani, Fragment C, pET۲۲b expression vector, pET۲۸a expression vector, E. coli BL۲۱ (DE۳) pLysS