Validation of mRNA/EST-based gene predictions in human Xp11.4 revealed differences to the organization of the orthologous mouse locus

Careful manual annotation of the human reference sequence provides a solid basis for the identification of disease-associated genes. Toward this end, we focused on a medically relevant 2.6-Mb region of the human chromosome Xp11.4 between markers DXS9851 and DXS9751 and identified 16 transcription units according to the Vertebrate Genome Annotation (Vega) rules. In order to validate these annotations, we performed a comprehensive RT-PCR expression analysis and a human-mouse comparison. This revealed, despite the high overall genomic conservation of the region, remarkable differences of the gene content between human and mouse. Whereas 12 of 16 annotations were confirmed by RT-PCR in human tissues, for only seven genes mouse orthologs could be identified and found to be expressed. This indicates that a comprehensive and experimentally supported annotation effort of the human genome simultaneously highlights regions with striking differences in gene organization to other species and may indicate evolutionary events specific to the human lineage demanding further functional analyses.