Cloning and expression of alkaline phosphatase gene from Schizosaccharomyces pombe

Abstract

A cDNA coding alkaline phosphatase (AP) homologue mas isolated from a cDNA library of Schizosaccharomyces pombe by colony hybridization. The nucleotide sequence of the cloned cDNA appeared to lack the N-terminal coding region, The genomic DNA encoding alkaline phosphatase homologue was isolated from S. pombe chromosomal DNA using PCR, The amplified DNA fragment was ligated into plasmid pRS315 to generate the recombinant plasmid pSW20. The DNA insert was subcloned as two smaller fragments for nucleotide sequencing. The sequence contains 2,789 bp and encodes a protein of 532 amino acids with a molecular mass of 58,666 daltons. The S, pombe cells containing plasmid pSW20 showed much higher AP activity compared with the yeast cells with vector only. This indicates that the cloned AP gene apparently encodes AP. The predicted amino acid sequence of the S, pombe AP shares homology with those of other known APs.