Study on the Ethnic Medicine Column

Genomic DNA extraction condition optimization of Arnebiae Radix based on response surface methodology

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  • 1. National Institutes for Food and Drug Control, Beijing 102629, China;
    2. China Pharmaceutical University, Nanjing 210009, China;
    3. Songyuan Institute for Food and Drug Control, Songyuan 138000, China

Online published: 2024-06-24

Abstract

Objective: To optimize the genomic DNA extraction conditions of Arnebiae Radix based on response surface methodology. Methods: The effect of different operation steps' reaction time in the plant genomic DNA extraction kit on the extraction efficiency and quality of Arnebiae Radix genomic DNA was investigated. The 3 conditions sampling amount, ethanol treatment, and lysis time in the Arnebiae Radix extraction process were optimized based on response surface method. The optimized conditions were tested and verified. The PCR amplification system of Arnebiae Radix genomic DNA was optimized, by adjusting the primer concentrations in the PCR amplification system. Results: The investigation result of different operation steps' reaction time in the plant genomic DNA extraction kit on the extraction efficiency and quality of Arnebiae Radix genomic DNA, showed that there was no obvious effect of different reaction time. Based on the response surface methodology, the optimal extraction conditions for the genomic DNA of Arnebiae Radix were determined as sampling amount of 100 mg, lysing overnight (12-15 h), and soaking the samples in 100% ethanol for 24 h for pretreatment. Through the verification of 100 batches of Arnebiae Radix market samples, the success rate of PCR amplification was 89%. The optimal primer concentration in the PCR system of the Arnebiae Radix genomic DNA was determined to be 0.10 μmol·L-1. If the genomic DNA still couldn't be amplified normally, the DNA can be diluted 10 times. Conclusion: The extraction and amplification efficiency of the Arnebiae Radix genomic DNA has been greatly improved, by optimizing the DNA extraction conditions and PCR amplification conditions. It provides support for the molecular biological identification for Arnebiae Radix. At the same time, it also provides a reference basis for the optimization of genomic DNA extraction conditions of other traditional Chinese medicine.

Cite this article

LIU Jie, FANG Wen-liang, TANG Zhe, LIAN Chao-jie, DAI Sheng-yun, GUO Li-nong, ZHENG Jian, QIAO Fei, MA Shuang-cheng, LI Yun-zheng . Genomic DNA extraction condition optimization of Arnebiae Radix based on response surface methodology[J]. Chinese Journal of Pharmaceutical Analysis, 2022 , 42(8) : 1345 -1353 . DOI: 10.16155/j.0254-1793.2022.08.07

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