Package and identification of replication-deficient recombinant adenovirus expression vector of human heparanase / 解放军医学杂志

ABSTRACT

Objective To construct a replication-deficient recombinant adenovirus expression vector of human heparanase (hpa). Methods The hpa gene was cloned at the downstream of CMV promoter of the adenoviral shuttle plasmid pDC315 in sense direction, and the resultant recombinant plasmid pDC315-hpa was transfected into HEK293 cell together with plasmid pBHGlox (deltaE1,3) containing adenoviral genome, then the replication-deficient recombinant adenovirus expression vector of hpa (Ad-hpa) was obtained, and it was identified by infection test, electronic microscope observation and PCR amplification. Results After purification and concintration,the titer of Ad-hpa reached 5?10 10pfu/ml. Virus particles could be found in virus concintration solution, and replication of virus was observed in HEK293 cells was observed under transmission electron microscope. Both adenovirus and hpa special fragment could be amplified from Ad-hpa by PCR, whereas hpa special fragment could not be amplified from the control. Conclusion The replication-deficient recombinant adenovirus expression vector of hpa was constructed successfully. This study established a foundation for further study on hpa vaccines and gene therapy for carcinoma.