Comparison of methods for DNA extraction from paraffin-embedded tissues and buccal cells.

Author(s): Cao W, Hashibe M, Rao JY, Morgenstern H, Zhang ZF

Publication: Cancer Detect Prev, 2003, Vol. 27, Page 397-404

PubMed ID: 14585327 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The purpose of this study was to determine the effects of DNA extraction method on DNA quality and PCR success from FFPE lung and bladder tumor specimens.

   Summary of Findings:

   The highest average DNA yield (48.6 ug) was obtained when DNA was extracted using the simple boiling method, but the OD 260/280 ratio was only 1.13. When DNA was extracted with phenol-chloroform and the QIAamp DNA mini kit, 6 and 9 ug of DNA were obtained, respectively, and the OD 260/280 ratios were 1.79 and 1.71, respectively. When DNA was extracted by phenol-chloroform or simple boiling, a 256 bp product of beta-globin was amplified in 29 and 30 of 34 specimens, respectively. In contrast, when DNA was extracted by the QIAamp DNA mini kit, beta-globin was successfully amplified in only 18 of 34 specimens.

   Biospecimens

   • Tissue - Lung
   • Tissue - Bladder

   Preservative Types

   • Formalin

   Diagnoses:

   • Neoplastic

   Platform:

   Analyte	Technology Platform
   DNA	PCR
   DNA	Spectrophotometry

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Analyte Extraction and Purification	Analyte isolation method	Phenol-chloroform Boiling method QIAamp DNA mini kit

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2. Study Purpose

   The purpose of this study was to determine the effects of DNA extraction method, storage and amplicon on PCR success from buccal cell specimens. After storage at 4 degrees C or room temperature, most specimens were frozen at -80 degrees C until analysis

   Summary of Findings:

   When DNA was extracted from buccal cells using the simple boiling method, beta-globin PCR was successful in only 12.5% of specimens. In contrast, beta-globin was successfully amplified in 94.1% and 75% of specimens extracted with phenol-chloroform and the QIAamp DNA mini kit, respectively. PCR success was not dependent on the targeted gene when DNA was extracted by phenol-chloroform or the QIAamp DNA mini kit; however, only beta-globin (256 bp) was successfully amplified from DNA extracted by the simple boiling method. The authors report that storage at room temperature or in the freezer had little effect on the ability to amplify DNA from buccal cells.

   Biospecimens

   • Cell - Buccal

   Preservative Types

   • None (Fresh)
   • Other Preservative
   • Frozen

   Diagnoses:

   • Not specified
   • Neoplastic

   Platform:

   Analyte	Technology Platform
   DNA	PCR
   DNA	Spectrophotometry

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Analyte Extraction and Purification	Analyte isolation method	Phenol-chloroform Boiling QIAamp DNA mini kit
   Storage	Storage duration	< 1 week > 1 week < 10 weeks > 10 weeks
   Storage	Storage temperature	Room temperature 4 degrees C -80 degrees C
   PCR Specific	Length of gene fragment	179 bp 219 bp 256 bp 459 bp
   PCR Specific	Targeted nucleic acid	GSTM1 Beta-globin GSTT1 GSTP1
   Biospecimen Preservation	Type of fixation/preservation	None (fresh) Frozen

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