摘要
目的分析产肠毒素大肠埃希菌stp毒力突变基因序列,及时优化监测方案,对食品中的产肠毒素大肠埃希菌进行有效的监测。方法利用生物信息软件对测序结果进行比对,分析变异位点。通过评估不同引物探针检测方案对性能的影响,确定最优的解决方案。结果通过基因序列测定,发现stp基因监测方案中上游引物第15号位置和16号位置发生了突变,NCBI中stp基因序列共63条,这条stp上游引物方案经过序列分析发现第15号位的T突变成A的共有12条,占比19%。第16号位的C突变成A的有11条,占比17%。优化后的stp检测方案能够正确扩增,除检出目标菌特异性条带无非特异条带产生,灵敏度达到0.05 ng/μl。结论优化后的检测方案准确度高,特异性好、灵敏度好,并且不干扰其他毒力基因的检测性能,满足食品中对产肠毒素大肠埃希菌的检测要求。
Objective To analyze the sequence ofstpvirulence gene of enterotoxigenicEscherichia coli,optimize the monitoring system,and effectively monitor enterotoxigenicEscherichia coliinfection in food.Methods The sequencing results were compared and the mutation sites were analyzed by bioinformatics software.The optimal solution is determined by evaluating the performance of different primer probe detection systems.Results It was found that there were mutations at position 15 and position 16 of the forward primer in thestpgene monitoring system.There were 63stpgene sequences in NCBI.Through sequence analysis,it was found that there were 12 T into A mutations at position 15,accounting for 19%.There are 11 cases in which C becomes A at position 16,accounting for 17%.The optimizedstpdetection system can amplify correctly the specific bands of the target bacteria,no non-specific bands are generated and the sensitivity can reach 0.05 ng/μl.Conclusion The optimized detection system has high accuracy,good specificity and sensitivity,and does not interfere with the detection performance of other virulence genes.This meets the requirements for the detection of enterotoxigenicEscherichia coliin food.
作者
刘俊华
吴玲玲
王雷
张广伟
杨海英
廖兴广
LIU Jun-hua;WU Ling-ling;WANG Lei;ZHANG Guang-wei;YANG Hai-ying;LIAO Xing-guang(Henan Provincial Center for Disease Control and Prevention,Henan Provincial Key Laboratory of Foodborne Disease Pathogen Tracing,Zhengzhou,Henan 450016,China;不详)
出处
《中国卫生检验杂志》
CAS
2022年第14期1681-1683,1690,共4页
Chinese Journal of Health Laboratory Technology
基金
河南省医学科技攻关计划项目(LHGJ20190690)。
