Isolation and Sequence Analysis of GpdII Promoter of the White Button Mushroom (Agaricus bisporus) from Strains Holland737 and IM008

Abstract

Many recent studies have shown that glycosylation patterns of <em>Agaricus bisporus</em> are similar to those of mammalians, so that this organism is a good candidate for the expression of glycosylated pharmaceutical protein. To achieve constant interested gene expression in all cells of the organism, proper promoter isolation is necessary. To isolate this promoter, PCR with specific primers was performed on extracted DNA of the white button mushroom strains Holland737 and IM008. The PCR amplified 290 bp fragments of gpdII promoters. IM008 gpdII promoter was used to construct pCAMBIAH8 plasmid. Comparison of isolated promoters among sequence records at NCBI demonstrated high similarity between IM008 gpdII promoter and previously reported gpdII promoter. Sequence analysis of isolated promoters revealed several point mutations on this promoter. TACAAA promoter sequence in −65 site acts as TATA box. Among the three CAAT candidate sequences, one is functional, which is located at position −108. Transformation of the white button mushroom with constructed pCAMBIAH8 plasmid was successfully performed.