Cloning and identification of human CYP2E1 cDNA and construction of its recombinant bacmid DNA for baculovirus expression system

Objective To obtain the recombinant bacmid w ith human cytochro me P 450 2E1(CY P2E1)cDNA suitable for ex pression in Bac-to-Bac baculovirus expression system.Methods Polymerase chain reaction(PCR)and DNA recom binant techniques were used in the experiments to obtain CYP2E1 cDNA from an unknown plasmid with CYP2E1 gene and clone it into pCMV-Mycv ector.The cDNA segment was analyzed by restriction enzyme digesting and DNA sequencing.Then the CYP2E1 cDNA was subcloned into vector pFastBac1,and was transformed into competent E.coli DH 10Bac,generating the recombinant bacmid.Results The gene segment was confirmed to be the whole coding region of CYP2E1 cDNA by DNA sequencing analysis and the recombinant bacmid was confirmed to have the whole 1.5kb of CYP2E1 cDN A.Conclusion The recombinant bacmid with CYP2E1 cDNA was constructed for making preparation for the next expression of CYP2E1 in Bac-to-Bac baculovirus expression system.