Cloning of the phytase gene phyA from Aspergillus ficuum 3.4322 and its expression in yeast

A phyA gene was cloned from Aspergillus ficuum 3.4322 by reverse transcription polymerase chain reaction. The amplified fragment was cloned into the pMD18 T-vector and sequenced. Nucleotide sequence analysis of the phyA gene showed that it comprised 1347bp without the signal peptide sequence and encodes a polypeptide of 448 amino acids. The phyA sequence has been deposited in GenBank (accession number: AF537344). Expression vectors pYPA1 and pYPA2 were constructed by cloning the phyA gene with and without the signal peptide sequence into the yeast expression vector pYES2. The recombinant plasmids were transformed into Saccharomyces cerevisiae INVSc1 by the method of LiAc. Phytase activity was found in pYPA2 (about 11.551U/ml) endocellular fluid and in pYPA1 supernatant (about 11.60IU/mL) by galactose inducing. The results demonstrated that the phyA gene had been expressed in Saccharomyces cerevisiae and the signal sequence of Aspergillu ficuum 3.4322 could facilitate the phytase secretion from S. cerevisiae efficiently.