2009 Vienna - European Society of Human Genetics

Genomics, Genomic technology and Epigenetics
4qA arrays with more than 10 repeat units, as well as 4qB and 10q
arrays, are non-pathogenic.
The current genetic diagnostics relies on Southern blotting, which allows
sizing of D4Z4 repeat arrays and, to some extent, determination
of their chromosomal origin and haplotype. However, it is cumbersome
and cannot reliably account for relatively frequent deviations from the
canonical description above, such as recombinations, translocations
between 4q and 10q, deletions of sequences adjacent to the repeat
arrays, somatic mosaicism etc. To bypass these heavy limitations, we
have developed a test based on a novel approach, the direct visualization
of the D4Z4 loci at kilobase-resolution on individual DNA molecules
stretched by Molecular Combing.
Our results on reference cell lines and patient samples demonstrate
the ability of Molecular Combing to precisely visualize the haplotype
and provide the repeat unit counts of all four 4q- and 10q- D4Z4 arrays
in a single experiment. Furthermore, we identified unexpected rearrangements
at the D4Z4 loci. This approach should allow for routine
genetic testing, as well as provide better understanding of the pathophysiology
of FSHD.
P11.038
A unique tool to detect LDLR, APOB and PcsK9 point mutations
as well as copy Number variation of the LDLR gene
M. Stef 1 , L. Palacios 1 , M. Viegas 1 , D. Tejedor 1 , A. Martinez 1 , M. Pocovi 2 , J.
Defesche 3 ;
1 Progenika Biopharma SA, Derio, Spain, 2 Department of Biochemistry, University
of Zaragoza, Zaragoza, Spain, 3 Department of Vascular Medicine, Academic
Medical Center, Amsterdam, The Netherlands.
Familial Hypercholesterolemia (FH) is an autosomal dominant disorder
with a prevalence of heterozygous FH of one in 500 individuals. It
is mostly due to point mutations in the low-density lipoprotein (LDL)receptor
gene (LDLR) and Copy Number Change (CNC) of part of this
gene account for 5-10%. Mutations are also seen in the APOB and
PCSK9 genes.
To improve FH genetic diagnosis, we have developed a new diagnostic
tool based on a DNA-array system: Lipochip which is able to detect at
the same time point mutations in the LDLR, APOB and PCSK9 genes
as well as CNC in the LDLR gene.
To enable detection of CNCs, controls probes from the chromosomes
X and 21 were included on the chip. 247 mutations were included on
the LIPOchip (238 LDLR, 3 APOB and 6 PCSK9 mutations). A software
was created for the automated analysis.
All point mutations and all types of CNC hybridized were identified.
Only one discrepancy was observed between MLPA and the LIPOchip,
due to a polymorphism in the sequence hybridizing the MLPA probe.
Reviewing the prevalence of the different mutations in Europe, we estimate
that this tool is able to detect more than 80% of the mutations of
representative European populations (Spain, Norway, Netherlands).
We here show that a unique tool is able to detect at the same time
point mutations and CNCs allowing rapid and robust genetic diagnosis
of FH.
P11.039
Analysis of the expression profile of mRNAs and microRNAs in
fragile X syndrome.
E. Tabolacci1 , M. Accadia1 , F. Pirozzi1 , M. Moscarda1 , U. Moscato2 , C. Bernardini3
, P. Chiurazzi1 , G. Neri1 ;
1 2 Institute of Medical Genetics, Rome, Italy, Institute of Public Health, Rome,
Italy, 3Institute of Human Anatomy, Rome, Italy.
Fragile X syndrome (FXS) is caused by the absence of the FMRP
protein. FMRP is involved in multiple pathways of mRNA metabolism
and stability [Zalfa et al., 2007]. Indeed, FMRP has been shown to be
associated to microRNAs (miRNAs) in mammals [Jin et al., 2004], possibly
regulating neuronal mRNA translation or stability.
We performed microarray experiments to characterize the transcriptional
profile of lymphoblastoid cell lines from 4 FXS, 3 controls and
2 unmethylated full mutation (UFM) carriers. Bioinformatic analysis
yielded a list of genes either up− or down−regulated in FXS lymphoblasts,
compared to controls. We selected 8 genes for validation and
confirmed that 5 of these were actually differentially expressed. We
also analyzed the miRNA expression profile in lymphoblasts from 6
FXS, 6 controls and 3 UFM carriers. We observed internal homogeneity
in each of the three groups (FXS, controls and UFM) and confirmed
the inter group difference (p