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目的:建立植物DNA混合物中物种成分实时荧光PCR定量方法。方法:建立PCR反应管中某物种DNA质量和该物种特异性基因扩增Ct值之间的定量关系,通过260 nm处光密度值的测定计算DNA提取液的浓度和DNA总质量,并以某物种DNA和总DNA的质量百分比表示该物种DNA含量。对56个待测样品中的物种成分进行定量测定以验证新建方法的准确性和精密度。结果:新建方法的LOQ低于0.51%;B ias为0.00%~20.79%,RSD为0.43%~19.64%。结论:新建方法的准确度和精密度均符合现行要求。
Objective: To establish a real-time fluorescence quantitative PCR method for species composition in plant DNA mixtures. Methods: The quantitative relationship between the DNA quality of a species in a PCR reaction tube and the Ct value of this species-specific gene amplification was established. The concentration of DNA extract and the total mass of DNA were calculated by measuring the optical density at 260 nm. The mass percentage of species DNA and total DNA indicates the DNA content of this species. The quantitative determination of species components in 56 samples to be tested to verify the accuracy and precision of the new method. Results: LOQ of the new method was lower than 0.51%, B ias was 0.00% ~ 20.79%, RSD was 0.43% ~ 19.64%. Conclusion: The accuracy and precision of the new method are in line with the current requirements.