MLPA-based approach for initial and simultaneous detection of GBA deletions and recombinant alleles in patients affected by Gaucher Disease.
The chromosomal region, in which the GBA gene is located, is structurally subject to misalignments, reciprocal and nonreciprocal homologous recombination events, leading to structural defects such as deletions, duplications and gene-pseudogene complex rearrangements causing Gaucher Disease (GD). Interestingly deletions and duplications, belonging to the heterogeneous group of structural defects collectively termed Copy Number Variations (CNVs), together with gene-pseudogene complex rearrangements represent the main cause of pitfalls in GD mutational analysis. In the present study, we set up and validate a Multiplex Ligation-dependent Probe Amplification (MLPA)-based approach to simultaneously investigate the potential occurrence of CNVs and complex rearrangements in 8 unrelated GD patients who had still not-well-characterized or uncharacterized alleles. The findings allowed us to complete the mutational analysis in 4 patients, identifying a rare deletion (g.-3100_+834del3934) and 2 novel recombinant alleles (g.4356_7031conJ03060.1:g.2544_4568; g.1942_7319conJ03060.1:g.1092_4856). These results demonstrate the diagnostic usefulness of MLPA in the detection of GBA deletions and recombinations. In addition, MLPA findings have also served as a basis for developing molecular approaches to precisely pinpoint the breakpoints and characterize the underlying mechanism of copy number variations.
Centro di Diagnostica Genetica e Biochimica delle Malattie Metaboliche, Istituto G. Gaslini, Via Gaslini 5, 16147 Genova, Italy. Electronic address: giulia.amico87@gmail.com.
Giulia
G
Amico
Centro di Diagnostica Genetica e Biochimica delle Malattie Metaboliche, Istituto G. Gaslini, Via Gaslini 5, 16147 Genova, Italy. Electronic address: serenagrossi@gaslini.org.
Serena
S
Grossi
MRC-Holland, Willem Schoutenstraat 1, 1057 DL Amsterdam, The Netherlands. Electronic address: r.vijzelaar@mlpa.com.
Raymon
R
Vijzelaar
Centro di Diagnostica Genetica e Biochimica delle Malattie Metaboliche, Istituto G. Gaslini, Via Gaslini 5, 16147 Genova, Italy. Electronic address: federicalanza@gaslini.org.
Federica
F
Lanza
Centro di Diagnostica Genetica e Biochimica delle Malattie Metaboliche, Istituto G. Gaslini, Via Gaslini 5, 16147 Genova, Italy. Electronic address: raffaellamazzotti@gaslini.org.
Raffaella
R
Mazzotti
Centro di Diagnostica Genetica e Biochimica delle Malattie Metaboliche, Istituto G. Gaslini, Via Gaslini 5, 16147 Genova, Italy. Electronic address: fabiocorsolini@gaslini.org.
Fabio
F
Corsolini
MRC-Holland, Willem Schoutenstraat 1, 1057 DL Amsterdam, The Netherlands. Electronic address: m.ketema@mlpa.com.
Mirjam
M
Ketema
Centro di Diagnostica Genetica e Biochimica delle Malattie Metaboliche, Istituto G. Gaslini, Via Gaslini 5, 16147 Genova, Italy. Electronic address: mirellafilocamo@gaslini.org.
Mirella
M
Filocamo
D000483:Alleles; D056915:DNA Copy Number Variations / Q000235:genetics*; D005260:Female; D005776:Gaucher Disease / Q000175:diagnosis / Q000235:genetics*; D020440:Gene Duplication / Q000235:genetics; D005962:Glucosylceramidase / Q000235:genetics*; D006801:Humans; D008297:Male; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D017384:Sequence Deletion / Q000235:genetics
D005962:Glucosylceramidase
Copy number variations; Deletions; GBA gene; Gaucher Disease; Gene-pseudogene complex rearrangements; Multiplex Ligation-dependent Probe Amplification (MLPA)