Studies of gene flow between closely related taxa can provide insight into the genetic basis of speciation. To evaluate the importance of the X chromosome in reproductive isolation between subspecies of the European rabbit and to study the genomic scale over which islands of differentiation extend, we resequenced a total of 34 loci distributed along the X chromosome and chromosome 14. Previous studies based on few markers suggested that loci in centromeric regions were highly differentiated between rabbit subspecies, whereas loci in telomeric regions were less differentiated. Here, we confirmed this finding but also discovered remarkable variation in levels of differentiation among loci, with FST values from nearly 0 to 1. Analyses using isolation-with-migration models suggest that this range appears to be largely explained by differential levels of gene flow among loci. The X chromosome was significantly more differentiated than the autosomes. On chromosome 14, differentiation decayed very rapidly at increasing distances from the centromere, but on the X chromosome distinct islands of differentiation encompassing several megabases were observed both at the centromeric region and along the chromosome arms. These findings support the idea that the X chromosome plays an important role in reproductive isolation between rabbit subspecies. These results also demonstrate the mosaic nature of the genome at species boundaries.