Cloning of differentially expressed genes in highly and low metastatic rat osteosarcomas by a modified cDNA-AFLP method

Tomokazu Fukuda, Akira Kido, Kazunori Kajino, Masahiro Tsutsumi, Yoshizumi Miyauchi, Toshifumi Tsujiuchi, Yoichi Konishi, Okio Hino

Research output: Contribution to journalArticlepeer-review

20 Citations (Scopus)

Abstract

To identify differentially expressed genes between highly and low metastatic rat transplantable osteosarcomas, we applied a modified AFLP (amplified fragment length polymorphisms) method for cDNA subtraction. The specific point of our modification is selective amplification using suppression PCR technique after restriction enzyme cutting. Our cDNA-AFLP gave high reproducibility (about 95%) in mRNA patterns and enabled us to clone four dominantly expressed genes in a highly metastatic tumor line. Three showed homology with known genes, encoding Ki-67, a proliferation-associated effective marker of malignancy, type IV collagen alpha-3, a major component of basement membrane, and KIAA77 for which the function is unknown. Although one fragment showed no database homology, we revealed a derivation from the rat homologue of the Drosophila melanogaster diaphanous gene (Dia) by cloning of longer cDNA. Dia genes, known to affect actin filament formation, are downstream effectors of Rho small GTPase. The results suggest that alterations in the expression of cytoskeletal protein, basement membrane elements, and proliferative markers may be important for metastasis of osteosarcomas.

Original languageEnglish
Pages (from-to)35-40
Number of pages6
JournalBiochemical and biophysical research communications
Volume261
Issue number1
DOIs
Publication statusPublished - 1999 Jul 22

Keywords

  • Metastasis
  • Osteosarcomas
  • Subtraction
  • cDNA-AFLP

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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