Efficiency of extraction and amplification of DNA from formalin-fixed paraffin-embedded archival bone marrow trephine specimens
Purpose: Formalin-fixed paraffin-embedded tissue is an invaluable source for retrospective molecular genetic studies, including gene clonality assays. It is however conventionally believed that extraction of high-quality genomic DNA may be problematic, especially from bone marrow trephine samples. The purpose of this study was to determine the efficiency of DNA extraction from archived bone marrow trephine specimens fixed in formalin.
Methods: DNA was extracted from formalin-fixed paraffin-embedded archived bone marrow trephine specimens using the Roche High Pure PCR Template Preparation Kit. Efficiency of DNA amplification at 84, 96, 200, 300, 400 and 600 base pairs (bp) was determined using a master mix (Specimen Control Size Ladder) that targets multiple genes from a gene clonality assay kit based on the Biomed-2 study.
Results: 136 samples from 128 formalin-fixed archived bone marrow trephines were used for the study. Positive amplification was noted in 117/136 specimens (86.02%) with a maximum number of samples amplifying at 96-200 bp. 108 samples (79.41%) demonstrated positive amplification at 96 bps and 67/136 (49.26%) at 200 bps. A progressive reduction in amplification was noted at higher bps with 34 positive cases (25.0%) at 300 bps, 27 (19.85%) at 400 bps and 18 (13.23%) at 600 bps.
Conclusions: Although it has been believed that DNA obtained from archival bone marrow trephine specimens is of poor quality, positive amplification was obtained in 86% of formalin-fixed paraffin-embedded samples in our study, indicating good potential for utilisation of PCR techniques for detection of gene clonality on bone marrow trephines.