食品检验用标准菌株分子水平质控方法的建立与应用

目的 建立食品检验用标准菌株分子水平质控方法，并进行应用。 方法 使用16S rRNA基因序列分析、脉冲场凝胶电泳（PFGE）、多位点序列分型等分子生物学技术手段，对食品安全国家标准中使用的标准菌株进行分子确认，并对不同的批号进行验证。 结果 16S rRNA基因序列比对结果符合该菌株所在的菌属，并获得标准菌株16S rRNA基因标准序列，不同批号的标准菌株16S rRNA基因序列完全一致；确定了每株标准菌株的ST型和基因型，并对不同批号的菌株进行了基因型比较，未发现型别改变；对无PulseNet标准操作方法的菌株，开展方法学研究，获得了最适用的限制性内切酶和电泳参数，建立了标准菌株分子指纹图谱，比较不同批号菌株的PFGE图谱，相似性均为100%。 结论 本研究首次将分子生物学技术应用到标准菌株的质量控制，突破了使用传统质控方法的瓶颈，从分子水平实现对标准菌株稳定性的评价，保证了标准菌株在食品检验过程中的稳定性和一致性。

Establishment and application of molecular quality control methods for standard strains for food inspection

Objective The molecular quality control methods were established and applied for the standard strains of food inspection. Methods The different molecular methods, such as 16S rRNAgene sequence analysis, pulsed-field gel electrophoresis (PFGE), and multi locus sequence typing, were used for the molecular confirmation of the standard strains of food inspection used according to national food safety standards, and they were verified by the different batch strains. Results The results obtained after 16S rRNA gene sequence alignment were consistent with the genus of the strain and the 16S rRNA gene sequences of different batch standard strains were identical; Sequence typing (ST) type of the standard strains were identified and the ST of different batch isolates were identical. The research on the method of PulseNet was exploded on the strains without the standard method to obtain the suitable condition of restrictive endonuclease and electrophoresis parameters. The methods were then used to establish molecular fingerprints of standard strains. The PFGE profile similarity of different batches was 100%. Conclusion The molecular biology technology was applied on the quality control of the standard strains for the first time in this study. It hit the bottleneck of traditional quality control methods. The stability of the standard strains was assessed at the molecular level to ensure the stability and consistence of the standard strains in the food inspection process.