Performance, protein synthesis and mucosal DNA in small intestine of Leghorn hens may be affected by low quality feedstuff. An experiment was conducted in completely randomized design (CRD) in 2 × 2 factorial arrangement. Main factors included diets containing 20 and 40 % barley and black and blue strains of leghorn. Layer performance was recorded weekly up to eight weeks. Egg production (%), egg mass and egg number were significantly higher in black strain leghorn on 20% barley diet than blue strain with 40% barley diet (P<0.05). Treatments blue × 40% barley and black × 20% barley showed lowest and highest egg production percentage, egg mass and egg number, respectively (P<0.05). The lowest protein synthesis in small intestine was shown in blue × 40% barley treatment group. Small intestine mucosal DNA quantity in blue strain was significantly higher than that in black strain (P<0.05). In contrast, protein synthesis in blue strain was lower than that in black strain (P<0.05). It was concluded that lower performance in blue × 40% treatment group could be related to lower protein synthesis in small intestine in blue strain, and this may be due to the high barley (40%) contained in diet.Key words: Performance, protein synthesis, mucosal DNA, small intestine