Expression of lignin peroxidase H2 from Phanerochaete chrysosporium by multi-copy recombinant Pichia strain

The lipH2 gene, encoding the expression of lignin peroxidase, was cloned from Phanerochaete chrysosporium BKM-F-1767 and expressed in Pichia pastoris X-33, a yeast. The cDNA of LiPH2 was generated from total RNA extracted from P. chrysosporium by PCR with primers that do not contain a P. chrysosporium lignin peroxidase secretion signal. The gene was then successfully inserted into the expression vector pPICZalpha, and resulted in the recombinant vector pPICZalpha-lipH2. The transformation was conducted in two ways. One was using the wild Pichia pastoris as the recipients, which results in the recombinant P. pastoris with single or low lipH2 gene copy. The second was using P. pastoris and single or low lipH2 gene copy as the recipients, which results in the recombinant P. pastoris with multi-copies of lipH2 genes. This study firstly expressed the gene lipH2 in P. pastoris and achieved the successful expression of the lipH2 depending upon the generation of a recombinant strain that contained multiple copies. The lignin peroxidase activity reached a maximum of 15 U/L after 12 h induction.