Detection of genetically modified soybean event DAS-44406-6 by real-time PCR method and digital PCR method

Purpose: To determine the flanking sequence between exogenous and endogenous fragments using rapid amplification of cDNA ends (5'-RACE) and consequently to examine genetically modified soybean (GMS) DAS-44406- 6. Methods: Specific primers and probes were designed based on the flanking sequences of exogenous fragments of DAS- 44406-6. The specificity of the developed method was validated by using it to detect a variety of other GM samples and non- GM samples. DAS-44406-6 was used to prepare 6 content gradients to test the sensitivity of the method. At last, digital PCR was applied to measure nucleic acid molecules for absolute quantification. Conclusions: A real-time PCR method has been established for the identification of GMS soybean DAS-44406-6 with high specificity. The limit of detection (LODs) of the real-time PCR method at template DNA concentration of 100 ng/reaction and 0.01% GM soybean content was 16.6 copies of genomic DNA from DAS-44406-6. As for the LOD of the digital PCR method, the relative standard deviation (RSD) was 0.7% at template DNA concentration of 0.5 ng/reaction. The highly specific method combining real-time PCR and digital PCR could meet the requirements for the detection of GMS DAS-44406-6. © 2016, China Food Publishing Company. All right reserved.