Reliable resequencing of the human dystrophin locus by universal long polymerase chain reaction and massive pyrosequencing (Articolo in rivista)

Type

• Articolo in rivista (Classe)
• Prodotto della ricerca (Classe)

Label

• Reliable resequencing of the human dystrophin locus by universal long polymerase chain reaction and massive pyrosequencing (Articolo in rivista) (literal)

Anno

• 2010-01-01T00:00:00+01:00 (literal)

Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#doi

• 10.1016/j.ab.2010.07.022 (literal)

Alternative label

• Bonnal RJ, Severgnini M, Castaldi A, Bordoni R, Iacono M, Trimarco A, Torella A, Piluso G, Aurino S, Condorelli G, De Bellis* G, Nigro V (2010)
  Reliable resequencing of the human dystrophin locus by universal long polymerase chain reaction and massive pyrosequencing
  in Analytical biochemistry (Print)
  (literal)

Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#autori

• Bonnal RJ, Severgnini M, Castaldi A, Bordoni R, Iacono M, Trimarco A, Torella A, Piluso G, Aurino S, Condorelli G, De Bellis* G, Nigro V (literal)

Pagina inizio

• 176 (literal)

Pagina fine

• 184 (literal)

Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#numeroVolume

• 406 (literal)

Rivista

• Analytical biochemistry (Rivista)

Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#numeroFascicolo

• 2 (literal)

Note

• ISI Web of Science (WOS) (literal)

Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#affiliazioni

• 1. CNR, ITB, I-20090 Segrate, Italy 2. Telethon Inst Genet & Med TIGEM, I-80131 Naples, Italy 3. Univ Naples 2, Med Genet Lab, Dipartimento Patol Gen, I-80138 Naples, Italy 4. IRCCS Multimed PST, I-20100 Milan, Italy (literal)

Titolo

• Reliable resequencing of the human dystrophin locus by universal long polymerase chain reaction and massive pyrosequencing (literal)

Abstract

• he X-linked dystrophin gene is well known for its involvement in Duchenne/Becker muscular dystrophies and for its exceptional megabase size. This locus at Xp21 is prone to frequent random molecular changes, including large deletions and duplications, but also smaller variations. To cope with such huge sequence analysis requirements in forthcoming diagnostic applications, we employed the power of the parallel 454 GS-FLX pyrosequencer to the dystrophin locus. We enriched the genomic region of interest by the robust amplification of 62 fragments under universal conditions by the long-PCR protocol yielding 244,707 bp of sequence. Pooled PCR products were fragmented and used for library preparation and DNA sequencing. To evaluate the entire procedure we analyzed four male DNA samples for sequence coverage and accuracy in DNA sequence variation and for any potential bias. We identified 562 known variations and 55 additional variants not yet reported, among which we detected a causative Arg1844Stop mutation in one sample. Sanger sequencing confirmed all changes. Unexpectedly, only 3x coverage was sufficient for 99.9993% accuracy. Our results show that long PCR combined to massive pyrosequencing is very reliable for the analysis of the biggest gene of the human genome and open the doors to other demanding applications in molecular diagnostics. (literal)

Prodotto di

• Tecnologie ad Array e di sequenziamento massivo (PM.P06.009.001) (Modulo)

Autore CNR

• MICHELE IACONO (Persona)
• RAOUL JEAN PIERRE BONNAL (Persona)
• ROBERTA BORDONI (Unità di personale interno)
• GIANLUCA DE BELLIS (Persona)
• MARCO SEVERGNINI (Unità di personale interno)

Incoming links:

Autore CNR di

• RAOUL JEAN PIERRE BONNAL (Persona)
• ROBERTA BORDONI (Unità di personale interno)
• GIANLUCA DE BELLIS (Persona)
• MARCO SEVERGNINI (Unità di personale interno)
• MICHELE IACONO (Persona)

Prodotto

• Tecnologie ad Array e di sequenziamento massivo (PM.P06.009.001) (Modulo)

Http://www.cnr.it/ontology/cnr/pubblicazioni.owl#rivistaDi

• Analytical biochemistry (Rivista)