International audienceThe vast majority of reported cases of resistance to ALS inhibitors in broadleaves is due to the selection of mutant, herbicide-resistant ALS alleles carrying a mutation at one of a few ALS codons that can be easily identified (ALS-based resistance). Non-ALS-based resistance (non-target-site resistance, NTSR), considered to be endowed by differences in the expression of many genes, has essentially been reported in grasses and hardly ever in broadleaves. We investigated NTSR to ALS inhibitors in poppy by pairing plants resistant to ALS inhibitors with susceptible plants and subsequently analysing the segregation of resistance in F1 families. Poppy plants resistant to ALS inhibitors were genotyped prior to pairing using a PCR-based assay and contained one (heterozygous mutant plants) or no mutant ALS allele. Pairings were made by hand-pollination. F1 progeny plants were genotyped at ALS following herbicide application to unambiguously link the phenotype (resistant or sensitive) to the ALS genotype (heterozygous mutant or non-mutant at ALS). F1 plants from pairings using a resistant parent plant wild-type at ALS were all non-mutant. Some of them were resistant to ALS inhibitors, as expected if NTSR is involved. Thus, NTSR to ALS inhibitors exists in poppy and is genetically inherited. F1 plants obtained from pairings using a resistant parent plant heterozygous mutant at ALS included more than 25% plants resistant to ALS inhibitor(s) that were non-mutant at ALS. This indicates that NTSR resistance to ALS inhibitors in Papaver rhoeas can occur alone, or together with the presence of mutant ALS alleles in individual plants. Segregation analysis of the F1 families also showed that the Mendelian segregation of mutant ALS alleles was not always respected. Our data indicate that the genetic control of resistance to ALS inhibitors in poppy is more complex than expected, and involves both ALS-based resistance and NTSR