Although pedigree selection is the most commonly used method for developing inbred lines of maize, there are no studies on its effect on the heterozygosity of the lines. The objective of this work was to study the effect of pedigree selection on their heterozygosity. Thirteen F5 or F6 maize inbred lines developed by the pedigree selection method in four breeding programs and their F1 and F2 − F4 ancestors were genotyped with simple sequence repeat markers distributed along the genome. Simulation was also conducted assuming different models of selection to investigate the selective forces needed to explain the data. In the F2, F3 and F4 40%, 66% and 86% of the markers segregating in the F1 were fixed; that is, in the F2 and F3 fixation was lower than neutral expectation, but higher in the F4. Due to such opposite apparent directions of selection, the heterozygosity of the lines in the F5 or F6 generations did not differ significantly from neutral expectations. The time to fixation differed from that expected with neutrality for most of the chromosomes, indicating that selection is distributed across the genome; but apparent overdominant effects in chromosome 7 were higher than in other chromosomes. In conclusion, the relationship between heterozygosity and vigour may reduce the effectiveness of pedigree selection in its goal of selecting the more vigorous, homozygous individuals. A more effective procedure is proposed using molecular markers for the identification of the more homozygous individuals, the most vigorous of those individuals being selected.