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Published on: August 24, 2017
A modified extraction method of circulating free DNA for epidermal growth factor receptor mutation analysis
Haihua Yuan1, Zhong-Zheng Zhu, Yachao Lu
1Department of Oncology, No. 3 People's Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai, China.
Yonsei Medical Journal
|December 22, 2011
Summary
An improved plasma cell-free DNA (cfDNA) extraction method using modified phenol-chloroform (MPC) yields more cfDNA and tumor DNA than standard kits. This enhances epidermal growth factor receptor (EGFR) mutation detection in non-small-cell lung cancer (NSCLC) patients.
Area of Science:
- Molecular Biology
- Oncology
- Biochemistry
Background:
- Circulating free DNA (cfDNA) in plasma shows promise as a non-invasive surrogate for tumor DNA.
- Current cfDNA extraction methods may be inefficient, leading to missed detection of epidermal growth factor receptor (EGFR) mutations.
Purpose of the Study:
- To develop and validate an efficient plasma cfDNA extraction protocol.
- To improve the detection of tumor-derived cfDNA for enhanced mutation analysis.
Main Methods:
- Comparison of a modified phenol-chloroform (MPC) method against a commercial kit (Qiagen) for plasma cfDNA extraction from non-small-cell lung cancer (NSCLC) patients.
- Quantification of cfDNA yield using TaqMan quantitative polymerase chain reaction (qPCR).
- Evaluation of extraction efficiency on EGFR mutation analysis using Mutant-enriched PCR (ME-PCR) sequencing and a commercial mutation test kit.
Main Results:
- The MPC method extracted significantly more plasma cfDNA than the Qiagen kit (p=0.011).
- A higher proportion of longer cfDNA fragments (≥ 202 bp) was observed with the MPC method (p=0.002).
- The MPC method demonstrated superior performance in detecting EGFR mutations and yielded more tumor-origin DNA.
Conclusions:
- The modified phenol-chloroform (MPC) method offers an improved and efficient approach for plasma cfDNA extraction.
- This enhanced cfDNA extraction protocol is beneficial for accurate EGFR mutation analysis in NSCLC patients.
