Efficient genetic analysis of microdissected samples by agarose-bead method: Alterations of β-catenin gene in fundic gland polyp and heterotopic gastric mucosa of duodenum

摘要

Molecular genetic analyses of archival formalin-fixed,and paraffin-embedded (FFPE) pathological specimens taken at biopsy or autopsy, are occasionally compromised because the DNA molecules therein are inevitably degraded. Furthermore, since these tissue samples comprise various cell types, the analyses based on mixtures of such heterogeneous populations often fail to reflect the nature of the affected cells. In the present study, to elucidate the contribution of β-catenin gene mutation to the fundic gland polyp and the heterotopic gastric mucosa in the duodenum, we successfully introduced an agarose-bead mediated technique as an effectual tool for retrospective morphology-oriented genetic analyses. Microdissected samples were embedded in low-melting agarose, and directly treated with proteinase K. A fragment of the agarose-bead was used as a template for polymerase chain reaction to analyze β--catenin mutation. Of the six cases of heterotopic gastric mucosa in the duodenum associated with fundic gland polyps, one showed a common 1-bp missense mutation at codon 37 shared by both the fundic gland polyp and the heterotopic gastric mucosa. Alternatively, a 1-bp silent mutation at codon 33 and missense mutation at codon 32 were identified only in the heterotopic gastric mucosa. Agarose-bead mediated technique shows superior sensitivity to the previously described techniques and is an effectual tool for retrospective morphologyoriented genetic analyses using a large number of archival pathological samples stored for long periods in the pathology laboratory.