Simple detection of genomic microdeletions and microduplications using QMPSF in patients with idiopathic mental retardation

Saugier-Veber, Pascale; Goldenberg, Alice; Drouin‐Garraud, Valérie; Rochebrochard, Céline de La; Layet, Valérie; Drouot, Nathalie; Meur, Nathalie Le; Gilbert‐Dussardier, Brigitte; Joly‐Helas, Géraldine; Moirot, H.; Rossi, Annick; Tosi, Mario; Frébourg, Thierry

doi:10.1038/sj.ejhg.5201661

Cited by 38 publications

(26 citation statements)

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“…QMPSF analysis has been shown to be a sensitive method for the detection of both deletions and duplications (Saugier-Veber et al, 2006 and is used in numerous molecular diagnostic laboratories. This approach allowed us to identify six intragenic OFD1 large deletions, spanning exons from 1 to 23 without hotspots.…”

Section: Discussionmentioning

confidence: 99%

Genomic deletions ofOFD1account for 23% of oral-facial-digital type 1 syndrome after negative DNA sequencing

et al. 2008

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Oral-facial-digital type I syndrome (OFDI) is characterised by an X-linked dominant mode of inheritance with lethality in males. Clinical features include facial dysmorphism with oral, dental and distal abnormalities, polycystic kidney disease and central nervous system malformations. Considerable allelic heterogeneity has been reported within the OFD1 gene, but DNA bi-directional sequencing of the exons and intron-exon boundaries of the OFD1 gene remains negative in more than 20 % of cases. We hypothesized that genomic rearrangements could account for the majority of the remaining undiagnosed cases. Thus, we took advantage of two independent available series of patients with OFDI syndrome and negative DNA bi-directional sequencing of the exons and intron-exon boundaries of the OFD1 gene from two different European labs: 13/36 cases from the French lab; 13/95 from the Italian lab. All patients were screened by a semiquantitative fluorescent multiplex method (QFMPSF) and relative quantification by real-time PCR (qPCR). Six OFD1 genomic deletions (exon 5, exons 1-8, exons 1-14, exons 10-11, exons 13-23 and exon 17) were identified, accounting for 5 % of OFDI patients and for 23 % of patients with negative mutation screening by DNA sequencing. INTRODUCTIONOral-facial-digital syndrome type I (OFDI; MIM# 311200) or Papillon-Léage-Psaume syndrome, which belongs to the heterogeneous group of oral-facial-digital syndromes (OFDS), is characterised by an X-linked dominant mode of inheritance and is lethal in males (Gorlin et al., 2001). Clinical features include facial dysmorphism with hypoplasia of alae nasi, oral frenula and clefts, lingual hamartoma, digital asymmetry with brachydactyly of hands, various degrees of mental deficiency, frequent brain malformations (absence of corpus callosum, porencephaly, hydrocephalus, vermis hypoplasia and Dandy-Walker malformation), and adult polycystic kidney disease (Scolari et al., 1997;Odent et al., 1998; Thauvin-Robinet et al., 2006, Prattichizzo et al., in press). Clinical variability is high and diagnosis may be difficult in minor cases. Moreover, the phenotype overlaps with those reported in the other forms of OFD syndromes (Gurrieri et al., 2007). The OFDI syndrome results from mutations in the OFD1 gene (MIM# 300170) (Xp22.2-22.3) with 23 coding exons (Ferrante et al., 2001). OFD1 is also implicated in a novel X-linked recessive mental retardation syndrome comprising macrocephaly and ciliary dysfunction (Budny et al., 2006). In OFDI syndrome, 92 mutations (frameshift, nonsense, missense and splice mutations) have been identified by DNA bi-directional sequencing of the exons and intron-exon boundaries of the OFD1 gene (Ferrante et al., 2001;Rakkolainen et al., 2002;Romio et al., 2003;Thauvin-Robinet et al., 2006; Prattichizzo et al., in press).To date, no mutation of the OFD1 gene has been detected in about 20% of patients presenting with clinical signs +/-a familial history consistent with OFD1 syndrome (Ferrante et al., 2001;Thauvin-Robinet et al., 2006; Prattichizzo et...

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Section: Discussionmentioning

confidence: 99%

Genomic deletions ofOFD1account for 23% of oral-facial-digital type 1 syndrome after negative DNA sequencing

et al. 2008

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“…18 We describe here a new assay for measuring SMN mRNA levels and compare the expression of the SMN2 gene in peripheral blood cells and in the skeletal muscle of SMA patients. This assay is inspired by the QMPSF method (quantitative multiplex PCR of short fluorescent fragments) that was developed for comparative quantitative analysis of genomic DNA, 19,20 but is applied here to reverse-transcribed mRNA.…”

Section: Introductionmentioning

confidence: 99%

A sensitive assay for measuring SMN mRNA levels in peripheral blood and in muscle samples of patients affected with spinal muscular atrophy

et al. 2007

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Different therapeutic strategies are currently evaluated in spinal muscular atrophy (SMA) that are aimed at increasing full-length (FL) mRNA levels produced from the SMN2 gene. Assays measuring SMN mRNA levels are needed. We have developed a sensitive, comparative assay based on multiplex fluorescent reverse-transcription polymerase chain reaction (RT -PCR) that can measure, in the same reaction, the levels of SMN mRNA with and without exon 7 sequences as well as those of total SMN mRNA. This assay allows to calculate directly the ratios of FL SMN mRNA to SMN mRNA without exon 7 (D7). We have used this assay to compare the levels of SMN transcripts in the blood of 75 unrelated normal subjects and of 48 SMA patients, and in muscle samples of 8 SMA patients. The SMN1 and the SMN2 genes produced very similar levels of total mRNA. Levels of transcripts lacking exon 7 were linearly dependent on the number of SMN2 copies, both in SMA patients and in controls. In patients, FL mRNA levels correlated with SMN2 copy number. A significant but weaker inverse correlation was also observed between FL or D7 mRNA levels and disease severity, but patients with three SMN2 copies and different SMA types displayed similar mRNA levels. A significantly higher FL to D7 ratio was measured in blood cells than in skeletal muscle (0.8070.18 versus 0.4770.11). This assay can be used as a sensitive biomarker for monitoring the effects of various drugs in forthcoming clinical trials of SMA.

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“…9,10 An important cause of apparently negative genetic testing could be large genomic rearrangements that are missed by direct sequencing and that usually account for 5 to 15% of the molecular defects responsible for autosomal recessive diseases. 11,12 Several techniques exist for detecting these large deletions or insertions, such as the multiplex ligation-dependent probe amplification (MLPA) assay 11,13 or the quantitative multiplex PCR of short fluorescent fragments (QMPSF), 14 previously used in our laboratory. 15 However, to date, a systematic screening for large genomic rearrangements at the SLC12A3 gene in GS is lacking, and only 1 16 of the 143 reported mutations in the Human Gene Mutation Database (http://www.hgmd.cf.ac.uk/ac/index.php) is a large genomic rearrangement.…”

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Spectrum of Mutations in Gitelman Syndrome

Vargas‐Poussou

Dahan

Kahila

et al. 2011

Journal of the American Society of Nephrology

199

194

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Gitelman's syndrome (GS) is a rare, autosomal recessive, salt-losing tubulopathy caused by mutations in the SLC12A3 gene, which encodes the thiazide-sensitive NaCl cotransporter (NCC). Because 18 to 40% of suspected GS patients carry only one SLC12A3 mutant allele, large genomic rearrangements may account for unidentified mutations. Here, we directly sequenced genomic DNA from a large cohort of 448 unrelated patients suspected of having GS. We found 172 distinct mutations, of which 100 were unreported previously. In 315 patients (70%), we identified two mutations; in 81 patients (18%), we identified one; and in 52 patients (12%), we did not detect a mutation. In 88 patients, we performed a search for large rearrangements by multiplex ligationdependent probe amplification (MLPA) and found nine deletions and two duplications in 24 of the 51 heterozygous patients. A second technique confirmed each rearrangement. Based on the breakpoints of seven deletions, nonallelic homologous recombination by Alu sequences and nonhomologous end-joining probably favor these intragenic deletions. In summary, missense mutations account for approximately 59% of the mutations in Gitelman's syndrome, and there is a predisposition to large rearrangements (6% of our cases) caused by the presence of repeated sequences within the SLC12A3 gene.

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Simple detection of genomic microdeletions and microduplications using QMPSF in patients with idiopathic mental retardation

Cited by 38 publications

References 35 publications

Genomic deletions ofOFD1account for 23% of oral-facial-digital type 1 syndrome after negative DNA sequencing

Genomic deletions ofOFD1account for 23% of oral-facial-digital type 1 syndrome after negative DNA sequencing

A sensitive assay for measuring SMN mRNA levels in peripheral blood and in muscle samples of patients affected with spinal muscular atrophy

Spectrum of Mutations in Gitelman Syndrome

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