Sequence analysis of filaggrin gene by novel shotgun method in Japanese atopic dermatitis

Takashi Sasaki, Jun Kudoh, Tamotsu Ebihara, Aiko Shiohama, Shuichi Asakawa, Atsushi Shimizu, Atsushi Takayanagi, Itaru Dekio, Chieko Sadahira, Masayuki Amagai, Nobuyoshi Shimizu

Research output: Contribution to journalArticlepeer-review

18 Citations (Scopus)


Background: Recent reports indicated that nonsense mutations in filaggrin (FLG) found in ichthyosis vulgaris (IV) patients are predisposing factors for atopic dermatitis (AD) with asthma. The exon 3 of FLG contains tandemly repeated, highly homologous, 11-13 sequence units of 972 or 975 bp, each of which corresponds to the coding sequence of the processed filaggrin with slight sequence difference. This unique gene structure has hampered the precise DNA sequence determination. Objective: We developed a novel DNA sequencing method "FLG-shotgun" to directly characterize the mutations in Japanese AD patients. Methods: We examined 24 Japanese AD patients with "FLG-shotgun" method. Results: Multiple units of FLG were amplified by PCR using several sets of common primers for the conserved regions, and DNA sequences of each cloned PCR product were determined. Multiple reads of DNA sequences in both alleles were aligned and re-constructed to cover the entire coding regions. We found three major genotypes (A, B, and C) which represent different numbers (11-13) of homologous sequence units. Furthermore, we found two novel nonsense mutations; one mutation 8666-8667CC>GA on the unit 9 of allele B that causes a nonsense mutation S2899X in two patients and the other mutation 9887C>A on the unit 10 of allele B that causes a nonsense mutation S3296X in two patients. Conclusion: We found two novel FLG mutations by directly analyzing Japanese patients with AD. FLG-shotgun will provide a valuable tool to further define the nature of the AD phenotype associated with FLG mutations.

Original languageEnglish
Pages (from-to)113-120
Number of pages8
JournalJournal of Dermatological Science
Issue number2
Publication statusPublished - 2008 Aug


  • Atopic dermatitis
  • DNA sequencing
  • Filaggrin
  • Shotgun

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Dermatology


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