Autosomal dominant ataxias (SCA1-2-3-6-7) and autosomal recessive Friedreich's ataxia (FRDA) are the most frequent ataxias in Europe. Mutations in SCA1-2-3-6-7 are CAG expansions that are transcribed and translated into a long sequence of polyglutamine in the corresponding proteins, resulting in a toxic function for the cell. PCRs were performed with pairs of specific primers for each SCAs, in which the forward primer was usually fluorescinated. PCR products were analyzed by gel electrophoresis and by capillary electrophoresis using ABI PRISM 3130xl gel Analyzer. Liz500 was used as internal molecular weight standard and GeneScan Software was used to elaborate the data. FRDA is a mitochondrial disease with a prevalence of 2x10-5 and a carrier frequency of 1/100. Almost 98% of the alleles has a pathological expansion of GAA in the first intron of the gene causing reduction of frataxin in homozygotes. Missense and truncating mutations are present in the remaining 2%. FRDA affected or carriers were identified by Hot-Start amplification using the XL-PCR kit (Applied Biosystems), with the following primers: GAA-F(5’GGG ATT GGT TGC CAG TGC TTA AAA GTT AG3’) GAA-R(5’GAT CTA AGG ACC ATC ATG GCC ACA CTT CGG3’). PCR products were analyzed on agarose gel and by capillary electrophoresis ABI PRISM 3130xl gel Analyzer. To confirm the diagnosis, Rox1000 and Liz500 were used as molecular weight standards and GeneScan Software was used to elaborate the data. 40 patients were analysed of which two resulted SCA2 affected, one was SCA1 affected, two were Freidreich affected and seven were Freidreich carriers.

Molecular diagnosis of dominant and recessive spinocerebellar ataxias

SELVATICI, Rita;FERLINI, Alessandra
2010

Abstract

Autosomal dominant ataxias (SCA1-2-3-6-7) and autosomal recessive Friedreich's ataxia (FRDA) are the most frequent ataxias in Europe. Mutations in SCA1-2-3-6-7 are CAG expansions that are transcribed and translated into a long sequence of polyglutamine in the corresponding proteins, resulting in a toxic function for the cell. PCRs were performed with pairs of specific primers for each SCAs, in which the forward primer was usually fluorescinated. PCR products were analyzed by gel electrophoresis and by capillary electrophoresis using ABI PRISM 3130xl gel Analyzer. Liz500 was used as internal molecular weight standard and GeneScan Software was used to elaborate the data. FRDA is a mitochondrial disease with a prevalence of 2x10-5 and a carrier frequency of 1/100. Almost 98% of the alleles has a pathological expansion of GAA in the first intron of the gene causing reduction of frataxin in homozygotes. Missense and truncating mutations are present in the remaining 2%. FRDA affected or carriers were identified by Hot-Start amplification using the XL-PCR kit (Applied Biosystems), with the following primers: GAA-F(5’GGG ATT GGT TGC CAG TGC TTA AAA GTT AG3’) GAA-R(5’GAT CTA AGG ACC ATC ATG GCC ACA CTT CGG3’). PCR products were analyzed on agarose gel and by capillary electrophoresis ABI PRISM 3130xl gel Analyzer. To confirm the diagnosis, Rox1000 and Liz500 were used as molecular weight standards and GeneScan Software was used to elaborate the data. 40 patients were analysed of which two resulted SCA2 affected, one was SCA1 affected, two were Freidreich affected and seven were Freidreich carriers.
File in questo prodotto:
Non ci sono file associati a questo prodotto.

I documenti in SFERA sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.

Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11392/1399131
 Attenzione

Attenzione! I dati visualizzati non sono stati sottoposti a validazione da parte dell'ateneo

Citazioni
  • ???jsp.display-item.citation.pmc??? ND
  • Scopus ND
  • ???jsp.display-item.citation.isi??? ND
social impact