A method for generating knock-out rodents including rats and mice is disclosed. The method involves mutagenizing a rodent with a mutagen, obtaining progeny of the mutagenized rodent, and identifying, among the progeny, one progeny that carries a loss-of-function modification of a target gene. The preferred mutagen for generating knock-out mice and rats is N-ethyl-N-nitrosourea (ENU). The preferred screening assays for identifying a progeny of a mutagenized animal that carries a loss-of-function modification are yeast truncation assays and yeast functional assays. Knock-out rodents generated by the method of the present invention are also within the scope of the invention.