Abstract
A girl presenting severe neonatal form of Marfan syndrome (MFS) was born to healthy parents. Mutational screening of patient’s DNA revealed an intronic change, 20 bp deletion in intron 34 (c.4211-32_-13del) in the fibrillin-1 gene (FBN1).
In silico predictions of the effect of the deletion on pre-mRNA splicing showed marked drop in both the 3’ splice site score and the polypyrimidine tract score. As we were not able to perform RT-PCR directly from patient’s RNA, we took advantage of splicing minigene assay.
We cloned exon 35 together with neighbouring intronic regions into the vector pET. After transfection of HeLa cell line with the wild type and mutant constructs, we isolated total RNA and performed RT-PCR.
Its results clearly showed that the mutation led to exon 35 skipping in the majority of RNA. In patient, this splicing aberration presumably caused the in frame deletion of 126 nucleotides from FBN1 mRNA.