SummarySupernatant malate dehydrogenase (MDH) isozymes (as visualized by starch gel electrophoresis) are encoded by two distinct gene loci in both the largemouth and smallmouth bass. When an interspecific F1hybrid is formed between these two fish, a unique MDH isozyme is generated. The results of freeze‐thaw molecular hybridization (which is the first application of this technique to MDH) indicate that this unique isozyme in the F1hybrid is a heterodimer composed of one subunit of each parental type. The F1hybrids produced F2hybrids which in turn formed the F3hybrid population. The inheritance of alleles at theMDH‐Blocus is consistent with a single Mendelian autosomal locus. Furthermore, there is no evidence of linkage between the lactate dehydrogenase‐E locus and theMDH