The isoamylase gene (ISO) of Pseudomonas amyloderamosa JD210, an isoamylase-hy roducin mutan was cloned in an isoamylase-deficient and transformable mutant strain K3l. By deletivn analysis, the ISO gene was found to be located within a 3.3 kilvbases Bamfll fragment. Ils nuclootide sequence contained an open reading franle of 2328 nucleotides (776 amino acids) encoding a secreted isvamylase precursor. TI1e ISO gene fragment was inserted into plasmids pKT230 and pBR 322 in opposite orientations. The expression of the ISO gene in the constructed plasntids was compared in P. _amyloderamosa K31, Pseudonzonas aeruginosa PAOI-161, Pseudomonas pufida mt-2 and Escherichia cali HBIOI. In all transfvrmed cells, the niajority of the isoamylase Qlduc_ed was secreted and hieher isoainylase activities were obtained in transformants with the transcriptional direction ul the IDU gene similu to the nearby drug-determinant gene of the vector.
