Reports of unacceptable shattercane (Sorghum bicolor) control with acetolactate synthase (ALS)-inhibiting herbicides prompted the investigation of 29 fields in central and south-central Nebraska for ALS-resistant (ALSr) shattercane. These fields were located in three distinct geographical areas designated C, G, and P. Shattercane from 13 fields spanning all three areas was resistant to 80 g ai ha−1 (2 X field rate) primisulfuron. Accessions C and G were more resistant than accession P to primisulfuron and nicosulfuron. Accessions C and G were susceptible to imazethapyr, whereas accession P was resistant. The ALS resistance was associated with alterations in the ALS enzyme. Primisulfuron I50 values for ALS from ROX (forage sorghum), C, G, and P were 7, 8,510, 8,870, and 714 nM, respectively; nicosulfuron I50 values were 647, 4,110, 4,070, and 1,460 nM, respectively; and imazethapyr I50 values were 5,440, 13,100, 11,800, and 51,700 nM, respectively. Based on cross-resistance and enzyme sensitivities, at least two biotypes are represented in the three accessions of ALSr shattercane. Shattercane individuals from accessions C, G, and P were intercrossed to determine if the ALSr genes in each of the accessions were at independent loci. All the F2 populations were resistant to 80 g ai ha−1 primisulfuron, suggesting that the ALSr alleles in the three accessions are at the same locus or possibly linked loci. When the C, G, and P accessions were crossed with the wild type (WT), comparisons between the F1, susceptible, and resistant populations showed that primisulfuron resistance was expressed as a dominant, partially dominant, and additive trait for the C, G, and P accessions, respectively. The differences in ALSr allelic interactions indicate that primisulfuron resistance developed independently in each of the three accessions.