Abstract: Abstract Objective: Try to clone the extracellular domain gene of human thyroid peroxidase(hTPO) and construct the baculovirus expression vector of it, so that we can carry out it’s expression in the baculovirus-insect cell expression system in the future. Method: Human TPO’s extracellular domain gene was obtained by PCR method. The fragment was ligated with pGEM3zf(+) vector and pFastBac1 vector sequently to generate recombinant hTPO-pGEM3zf(+) and hTPO-pFastBAC1. The hTPO-pFastBAC1 was used to transfect E.coli DH10Bac so as to get the recombinant baculovirus expression vector(hTPO-Bacmid). For each step, we identify the accuracy by several methods. Results: By methods of PCR, restriction endonuclease analysis and DNA sequencing, the recombinant hTPO-pGEM3zf(+),hTPO-pFastBAC1 and hTPO-Bacmid was confirmed the same as predicted. Conclusion: We have cloned hTPO’s extracellular domain gene successfully, and then generated recombinant hTPO-pGEM3zf(+) and hTPO-pFastBAC1 accurately. The latter one achieved recombination with Bacmid by site-specific transposition. So the recombinant baculovirus expression vector of hTPO was obtained. This work has established a basis for hTPO gene’s eukaryotic expression in insect cells.

Key words: human thyroid peroxidase, extracellular domain gene, PCR, baculovirus expression vector system