RT-PCR splicing analysis of the NF1 open reading frame

Hum Genet. 2002 May;110(5):495-502. doi: 10.1007/s00439-002-0714-6. Epub 2002 Apr 4.


Neurofibromatosis 1 (NF1) is an autosomal dominant condition whose molecular diagnosis is challenging because of the large size of the gene and the vast number of unique NF1 gene mutations. Some splicing and nonsense mutations have been shown to cause exon skipping. Recently, temperature-induced abnormal splicing has been found in NF1 in ex-vivo tissues. This prompted us to investigate the entire NF1 transcript for such aberrant splicing. We found several novel exon skips that appeared de novo or were present initially and increased in aged/cooled blood: exon 20, exons 20 and 21 combined, exon 33, exon 34, exon 37, exon 40, exon 45, exons 43 and 45 combined, part of exon 43, and the first codon of exon 12b. Some aberrant splice forms were undetectable when blood was drawn into Qiagen PAXgene tubes, rather than EDTA vacutainers, and we demonstrate how these aberrant splicing events are a potential pitfall for RNA-based NF1 mutation characterization. The same reverse transcription/polymerase chain reaction strategy was used to screen for novel NF1 alternative splicing in Schwann cells and seven other tissues. Even though no Schwann-specific alternative exons were identified, we found minor novel splicing isoforms differentially expressed such as skips of exon 37 and exon 40. Skipping of exon 43, part of exon 43, and the first codon of exon 12b were found in all tissues analyzed. These forms suggest greater tissue-based variability in the NF1 message than was previously thought and may indicate minor amounts of heterogeneity at the protein level.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Alternative Splicing / genetics*
  • Exons / genetics
  • Humans
  • Mutation / genetics
  • Neurofibromin 1 / genetics*
  • Open Reading Frames / genetics*
  • Organ Specificity
  • Polymorphism, Single-Stranded Conformational
  • Protein Isoforms / genetics
  • Reverse Transcriptase Polymerase Chain Reaction
  • Schwann Cells / metabolism
  • Temperature
  • Time Factors


  • Neurofibromin 1
  • Protein Isoforms