Rapid and practical detection of β-globin mutation causing β-thalassemia by fluorescence-based PCR-single-stranded conformation polymorphism analysis

Asako Takahashi-Fujii, Yoshizumi Ishino, Ikunoshin Kato, Yasuyuki Fukumaki

研究成果: Contribution to journalArticle査読

16 被引用数 (Scopus)

抄録

We report a useful method for daily clinical examination for the diagnosis of thalassemia. We applied a fluorescence-based image analyser to a non-radioisotopic PCR-single-stranded conformation polymorphism (SSCP) analysis to detect mutations in the β-globin gene. PCR primers were labelled with rhodamine X and the amplified fragments from the β-globin gene were resolved by non-denaturing polyacrylamide gel electrophoresis. After loading, the glass plate was set in the image analyser and scanned with a green laser. We detected four common mutations in exon I and two major mutations in intron 1 of the β-globin gene isolated from patients with β-thalassemia. Moreover, to discriminate mutations and natural polymorphisms, we used primers including one base mismatch at the polymorphic site, which can substitute the polymorphic site by a constant base in the PCR amplified fragment. This fluorescence-based system was simple to operate and results were obtained rapidly as clear image data. Therefore, once the optimal conditions of the electrophoresis are determined, this system will be suitable for daily clinical use, especially for screening of molecular defects and for the prenatal diagnosis of genetic disorders.

本文言語英語
ページ(範囲)385-393
ページ数9
ジャーナルMolecular and Cellular Probes
8
5
DOI
出版ステータス出版済み - 10 1 1994

All Science Journal Classification (ASJC) codes

  • 分子生物学
  • 細胞生物学

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