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Summary We report on a complete genome scan for quantitative trait loci (QTL) affecting milk protein percentage (PP) in the Italian Holstein-Friesian cattle population, applying a selective DNA pooling strategy in a daughter design. Ten Holstein-Friesian sires were chosen, and for each sire, about 200 daughters, each from the high and low tails of estimated breeding value for PP, were used to construct milk DNA pools. Sires and pools were genotyped for 181 dinucleotide microsatellites covering all cattle autosomes. Sire marker allele frequencies in the pools were obtained by shadow correction of peak height in the electropherograms. After quality control, pool data from eight sires were used for all subsequent analyses. The QTL heterozygosity estimate was lower than that of similar studies in other cattle populations. Multiple marker mapping identified 19 QTL located on 14 chromosomes (BTA1, 2, 3, 4, 5, 6, 8, 9, 12, 14, 17, 20, 23 and 27). The sires were also genotyped for seven polymorphic sites in six candidate genes (ABCG2, SPP1, casein kappa, DGAT1, GHR and PRLR) located within QTL regions of BTA6, 14 and 20 found in this study. The results confirmed or excluded the involvement of some of the analysed markers as the causative polymorphic sites of the identified QTL. The QTL identified, combined with genotype data of these candidate genes, will help to identify other quantitative trait genes and clarify the complex QTL patterns observed for a few chromosomes. Overall, the results are consistent with the Italian Holstein population having been under long-term selection for high PP.