Isozyme methodology was used to develop genetic markers for the Asparagus officinalis nuclear genome. Thirty-four enzymes were evaluated for potential expression of polymorphisms, of which 13 had sufficient staining activity and resolution in the gels for interpretation. Five of the 13 were polymorphic, including aconitase (ACO), endopeptidase (END), malate dehydrogenase (MDH), phosphoglucomutase (PGM), and shikimate dehydrogenase (SKD). Segregation data for some allozymes of the Aco-1, End-1, Mdh-1, Pgm-1, and Skd-1 loci are presented. Cosegregation data indicated 4.1% linkage between the Skd-1 and Pgm-1 loci. Isozyme analysis of A. densiflorus 'Sprengeri' revealed patterns different from A. officinalis for aspartate aminotransaminase (AAT), END, and triosephosphate isomerase (TPI), all of which have potential for use in selection and verification of cell fusion products between the two species.