 Hello, my name is Elizabeth Chilis and I am a PhD student at the University of Antwerp, more specifically at the Centre for Medical Genetics. In this video I will explain more about the deep intramarital mutation causing Marfan syndrome. Years ago, a large family came to clinic showing symptoms closely related to Marfan syndrome, an autosomal dominant connective tissue disorder. The family presented with skeletal and cardiovascular features, including turquic aortic aneurysm, tall stature and scoliosis. Marfan syndrome is caused by mutations in the fibrillin 1 gene, encoding for the fibrillin protein which is important in the extracellular matrix. First, denaturing pressure-liquid chromatography was used to identify the disease causing mutation in the fibrillin 1 gene, but none was found. Without any result, auto-diagnostic tests were performed, including signer sequencing, multiplexed ligation-dependent probe amplification and CNP analysis. Additional signer sequencing of other known aneurysm genes, like TGFB receptor 1 and 2, SMET3 and ACTED2, could also not explain the phenotype in the family. We then performed linkage analysis on 11 members of the family. A large link region was found on chromosome 15. This region included 80 genes with more than 15 candidate genes. To screen all of these genes we used whole exome sequencing, but again we did not find any disease causing mutations. As fbn1 was also located in the linked region, we decided to screen fbn1 again, but this time at mRNA level. For that purpose, we isolated mRNA from patient skin fibrovis, which were cultured in the presence or absence of pyromycin. This is an inhibitor of non-sosmediated mRNA decay. The mRNA was transcribed to cDNA and signer sequencing was performed. On the edge between exome 56 and 57, a double peak pattern was observed, which suggested the existence of a pseudo-acton, which was confirmed by purification of the aberrant PCR fragment. Sequencing of the intron revealed an 8-OHG transition, causing the generation of a new splice donor site. Together with an existing cryptic splice acceptor site, this resulted into the inclusion of a pseudo-acton of 90 base pairs. This pseudo-acton contains a stop codon. We could demonstrate that this aberrant mRNA is degraded by a non-sosmediated mRNA decay because it was only observed in the presence of pyromycin. The mutations segregated in the family, including a patient with borderline clinical features. Although more than 90% of FBM-1 mutations can be identified with the current diagnostic tools, deep hydraulic mutation will still be missed. These require cDNA sequencing or whole gene sequencing to be identified. For more information, you can find the article on human mutation. Finally, I'd like to thank all patients and all my co-authors.