基于核酸功能化的氧化石墨烯技术快速检测含SSeC基因鼠伤寒沙门氏菌的研究

Establishment of a new method based on nucleic acid functionalized GO for the rapid detection of Salmonella typhimurium carrying SSeC gene

目的：探索建立简单、高效、低成本的检测含SSeC基因鼠伤寒沙门氏菌的新方法。方法利用羧基荧光素（ FAM）修饰的核酸探针（ FAM-P）与氧化石墨烯（ GO）非共价偶联成功构建了FAM-P/GO碳纳米生物传感器。考察该体系对不同浓度靶序列基因和含该靶基因的不同浓度菌体样品检测的灵敏性,并通过碱基错配试验和种属特异性试验验证其特异性。结果实验表明,该纳米生物传感器对靶标序列SSeC基因检测的最低下限为0.05μmol/L,而且在0.05-1.0μmol/L范围内呈现良好的线性关系（R2=0.9921）,对目标菌检测的最低下限为103 CFU/ml,在103-108 CFU/ml范围内呈良好的线性关系（R2=0.9987）,此外,该方法对实验组的信噪比要远远大于对照组,因此具有很高的特异性。结论成功构建了FAM-P/GO碳纳米生物传感器,为含SSeC基因鼠伤寒沙门氏菌的快速检测提供了一种可能的新方法。

Objective To establish a simple, efficient and low-cost method for the detection of Salmonella typhimurium carrying SSeC gene. Methods In this study, a nano-biosensor （ FAM-P/GO） was successfully established based on the noncovalent assembly of carboxy-fluorescein （ FAM）-labeled probe and graphene oxide （ GO） . The target gene at different concentrations and SSeC gene-harbored bacterium sam-ples were detected by the FAM-P/GO nano-biosensor to evaluate its sensitivity. The specificity of the estab-lished nano-biosensor was evaluated by using DNAs with mismatched base pairs and single-stranded DNAs （ ssDNAs） extracted from various species. Results The established strategy for SSeC gene detection showed a good linear range of 0. 05-1. 0 μmol/L （R2=0. 992 1） with a lower limit of 0. 05 μmol/L. Moreover, the lower detection limit for target bacterium samples was 103 CFU/ml and the fluorescence intensity increased linearly with the concentration from 103 CFU/ml to 108 CFU/ml. The signal-to-noise （ S/N） of the experi-mental group was much greater than that of the control group, which indicated that the establish method was highly specific. Conclusion The FAM-P/GO nano-biosensor was successfully established in this study, which provided a new and possible way for the rapid detection of Salmonella typhimurium harboring SSeC gene.