Clomazone is a herbicide used in the cultivation of numerous crops due to its unique site of action and effectiveness on weeds. The differences in clomazone susceptibility among plants have been attributed to the differences in their complex clomazone metabolic pathways that are not fully understood. We previously identified two CYP81A cytochrome P450 monooxygenases that metabolize five chemically unrelated herbicides in multiple-herbicide resistant Echinochloa phyllopogon. Since the resistant E. phyllopogon have decreased clomazone susceptibility, involvement of these P450s in clomazone resistance was suggested. In this study, we revealed that each P450 gene endowed Arabidopsis thaliana (Arabidopsis) with clomazone resistance. Consistent with this, clomazone resistance co-segregated with resistance to other herbicides in F progenies of crosses between susceptible and resistant E. phyllopogon, suggesting that the P450s are involved in differential clomazone susceptibility in E. phyllopogon. Arabidopsis transformations of the other seven CYP81As of E. phyllopogon found that two more genes, CYP81A15 and CYP81A24, decreased Arabidopsis susceptibility to clomazone. Differences in substrate preference between clomazone and a herbicide that inhibits acetolactate synthase were suggested among the four CYP81A P450s. This study provides insights into clomazone metabolism in plants.