A novel approach for extracting DNA from formalin-fixed paraffin-embedded tissue using microwave

Abstract

Background: Formalin-fixed paraffin-embedded (FFPE) tissue is the gold standard procedure for tissue preservation. However, the extraction of DNA is a cumbersome procedure as the extracted DNA is fragmented because of the cross-linking effect of formalin. Hence, the aim of the present study is to extract DNA from FFPE tissues using different techniques with a specific objective of comparing the extracted DNA, both quantitatively and qualitatively.

Method: Ten samples of FFPE tissues were retrieved from the archives of the Department of Oral Pathology. Total genomic DNA was extracted by different methods which included QIAamp DNA FFPE Tissue Kit, Norgen DNA FFPE Tissue Kit, phenol-chloroform method, mineral oil-based extraction, M/10 NaOH solution method, and microwave method. A 280-base pair sequence was selected for evaluation of downstream amplification.

Results: The statistical analysis was performed using unpaired student's t-test to compare the DNA yields and quality obtained by microwave methods with other methods using SPSS software. Total genomic DNA retrieved by the microwave method was superior to other methods both qualitatively and quantitatively.

Conclusion: DNA extraction from FFPE tissues is an onerous task as irreversible bonds form between the nucleic acid during fixation which are difficult to break during DNA retrieval. Hence, the microwave method provides good total genomic DNA which gives better downstream results when compared with other methods.

Keywords: DNA extraction; Formalin-fixed paraffin-embedded; Microwave; Polymerase Chain Reaction (PCR); Tissue preservation.

Fig. 1

Gel image of extracted total genomic DNA by different methods, lane 1–2: microwave method; lane 3–4: QIAamp DNA FFPE Tissue Kit; lane 5–6: Norgen DNA FFPE Tissue Kit; lane 7–8: PC method; lane 9–10: mineral oil; lane 11–12: M/10NaOH; lane 15: DNA ladder 100–3000 bp. PC, phenol–chloroform; FFPE, formalin-fixed paraffin-embedded.

Fig. 2

Gel image of PCR product of β actin, Product size: 280bp, lane 1: 100-1500bp ladder, lane 2–3: microwave method; lane 4: QIAamp DNA FFPE Tissue Kit, lane 5: Norgen DNA FFPE Tissue Kit; lane 6: PC method; lane 7: mineral oil; lane 8: M/10 NaOH. PC, phenol–chloroform; FFPE, formalin-fixed paraffin-embedded.