One of the future issues in the forensic application of the haploid Y-chromosome (Y-chr) is surveying the distribution of the Y-chr variation on a micro-geographical scale. Studies on such a scale require observing Y-chr variation on a high resolution, high sampling efforts and reliable genealogical data of all DNA-donors. In the current study we optimised this framework by surveying the micro-geographical distribution of the Y-chr variation in the central-western European region named Brabant. The Duchy of Brabant was a historical region in the Low Countries containing three contemporary Belgian provinces and one Dutch province (Noord-Brabant). 477 males from five a priori defined regions within Brabant were selected based on their genealogical ancestry (known pedigree at least before 1800). The Y-haplotypes were determined based on 37 Y-STR loci and the finest possible level of substructuring was defined according to the latest published Y-chr phylogenetic tree. In total, eight Y-haplogroups and 32 different subhaplogroups were observed, whereby 70% of all participants belonged to only four subhaplogroups: R1b1b2a1 (R-U106), R1b1b2a2* (R-P312*), R1b1b2a2g (R-U152) and I1* (I-M253*). Significant micro-geographical differentiation within Brabant was detected between the Dutch (Noord-Brabant) vs. the Flemish regions based on the differences in (sub)haplogroup frequencies but not based on Y-STR variation within the main subhaplogroups. A clear gradient was found with higher frequencies of R1b1b2 (R-M269) chromosomes in the northern vs. southern regions, mainly related to a trend in the frequency of R1b1b2a1 (R-U106).

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