1. DNA insertions in the 5′-flanking region of the insulin gene have been studied by Southern-blot-hybridization techniques using a cloned gene probe in 159 unrelated Caucasians. 2. Subjects homozygous for a large DNA insertion at this locus were found to have a higher risk of concurrence of non-insulin-dependent diabetes (NIDD) than controls ( P < 0.01). 3. Analysis of a single large NIDD pedigree ( n = 67 in four generations; 11 diabetic subjects) showed no linkage between the size of DNA inserts and diabetes (Lod score = −5.7), suggesting a separate diabetogenic locus (or loci) in this family.