Biotechnology Bulletin ›› 2021, Vol. 37 ›› Issue (6): 286-294.doi: 10.13560/j.cnki.biotech.bull.1985.2020-1587

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Genomic DNA Rapid Preparation and PCR Detection Methods for Genetically Modified Papaya

PAN Zhi-wen(), CHEN Wei-ting, GAO Jie-er, ZHOU Feng, YAO Juan, WANG Sheng-bin, JIANG Da-gang()   

  1. Inspection and Testing Center for Ecological and Environmental Risk Assessment of Plant and Plant-Related Microorganism(Guangzhou),Ministry of Agriculture and Rural Affairs/College of Life Sciences,South China Agricultural University,Guangzhou 510642
  • Received:2020-12-30 Online:2021-06-26 Published:2021-07-08
  • Contact: JIANG Da-gang E-mail:panzhiwen@scau.edu.cn;dagangj@scau.edu.cn

Abstract:

The preparation of genomic DNA template is the key step in the detection of genetically modified papaya. In order to improve detection efficiency,it is very important to establish rapid genomic DNA preparation and PCR detection methods for papaya samples. In this study,a rapid preparation method of papaya genomic DNA was developed,including sample preparation,homogenisation with preparation buffer and supernatant dilution. After the rapid preparation of genomic DNA solutions from different papaya samples,the obtained genomic DNA solutions were verified by PCR method. The results of qualitative PCR showed that the amplified bands were clear when the genomic DNA solutions of the leaves and fruits were diluted for 5 times. The PCR amplification results of leaf genomic DNA were better than that of the fruits when the DNA solution were diluted for 25 times. The PCR amplification efficiencies of leaf genomic DNA were reasonable in the real-time fluorescence PCR reaction. Sensitivity testing results indicated the specific fragments of endogenous reference and exogenous genes were amplified as expected by both qualitative and real-time fluorescence PCR methods when the contents in the leaves and fruits samples was 0.1%. The rapid preparation and PCR detection methods of papaya genomic DNA established here is efficient and stable.

Key words: genetically modified papaya, DNA preparation, PCR, detection method