Construction of eukaryotic expression vector of recombinant immunotoxin rat IP-10-PE38KDEL and its expression in NIH 3T3 cells

Objective To construct a new recombinant immunotoxin expression vector by fusing rat IFN-γ-inducible protein gene (rIP10) and a truncated pseudomonas exotoxin A ramification (PE38KDEL) gene, and explore the expression of the rIP-10-PE38KDEL fusion protein in NIH 3T3 cells. Methods rIP-10 was cloned by polymerase chain reaction (PCR). PE38KDEL gene was gained from an prokaryotic expression vector plasmid PRKL459K-IL-4-PE38KDEL by restriction endonuclease digestion, and then inserted to the eukaryotic expression vector pcDNA3.1(+). After the eukaryotic recombinant vector pcDNA3.1(+)-rIP-10-PE38KDEL was identified by PCR, restriction endonuclease digestion, and sequence analysis, then the vector was transfected into NIH 3T3 cells by liposome protocol. Immunofluorescence method was used to confirm the expression of the fusion gene in the NIH 3T3 cells. Results PCR, restriction endonuclease digestion, and sequence analysis revealed the rIP-10-PE38PE38KDEL fusion gene was cloned into the eukaryotic expression plasmid vector pcDNA3.1(+) successfully. The pcDNA3.1(+)-rIP-10-PE38PE38KDEL fusion gene could express in the NIH 3T3 cell. Conclusion The result provide the basis for research of the targeted cytotoxic activity to Th1 cell and may have some potential value in clinical application.