目的 比较不同的双歧杆菌标准品制备法，筛选适用于实时荧光定量PCR检测的双歧杆菌标准品。方法 以双歧杆菌标准菌株作为实验菌株，分别采用菌株DNA法、PCR产物扩增纯化法、质粒DNA法制备梯度浓度标准品，运用紫外分光光度计和实时荧光定量PCR 技术进行检测，并对数据进行统计学处理。结果 实时荧光定量PCR检测发现，3种不同的双歧杆菌标准品制备法的标准曲线均 r 2 ＞0.990；不同制备法的双歧杆菌标准品DNA浓度和纯度检测比较，质粒DNA制备法所制的双歧杆菌标准品浓度和纯度较另外两种方法高，差异有统计学意义（ P<0.01）。结论 双歧杆菌质粒DNA制备法制备的标准品质量较高，适用于实时荧光定量PCR检测，可以为双歧杆菌分子生物学检测提供参考依据。
英文摘要:
Objective To compare different preparation methods for quantitative real-time PCR (qPCR) detection of Bifidobacteria. Methods Standard strains of Bifidobacteria were prepared with concentration gradients using strain DNA, PCR product amplification and purification, and plasmid DNA methods. The concentrations of Bifidobacteria were determined with ultraviolet spectrophotometer and real-time quantitative PCR. Results Greater than 0.99 R 2 in values of standard curves were achieved by all three preparation methods. The plasmid DNA method obtained a higher level of concentration and purity of Bifidobacteria than the other two methods （ P<0.01）. Conclusion The plasmid DNA method produces high quality preparations and is more suitable for real-time quantitative PCR, which can provide a reference for the molecular biological detection of Bifidobacteria.
