实时荧光跨越式滚环等温扩增技术检测蜂蜜掺伪大米糖浆  

Detection of Honey Adulterated Rice Syrup by Real-time Fluorescence SpanningRolling Circle Isothermal Amplification

作　　者：杨浩 徐慧 李欣妍 卢鑫[2] 杨倩 张伟[1,3,4] Yang Hao;Xu Hui;Li Xinyan;Lu Xin;Yang Qian;Zhang Wei(College of Food Science and Technology,Hebei Agricultural University,Baoding 071000,Hebei;Department of Science and Engineering,Hebei Agricultural University,Cangzhou 061100,Hebei;College of Life Sciences,Hebei Agricultural University,Baoding 071000,Hebei;Key Laboratory of Analysis and Prevention and Control of Pathogenic Microorganisms Caused by Zootopia in Hebei Province,Baoding 071000,Hebei;School of Public Health,Hebei University,Baoding 071000,Hebei)

机构地区：[1]河北农业大学食品科技学院,河北保定071000 [2]河北农业大学理工系,河北沧州061100 [3]河北农业大学生命科学学院,河北保定071000 [4]河北省人畜共患病原微生物分析与防控重点实验室,河北保定071000 [5]河北大学公共卫生学院,河北保定071000

出　　处：《中国食品学报》2024年第1期242-251,共10页Journal of Chinese Institute Of Food Science and Technology

基　　金：国家自然科学基金项目(32172288,31371772);河北省自然科学基金重点项目(C2019204342);河北省自然科学基金项目(C2019204284);中央引导地方科技发展资金项目(216Z5501G,226Z5503G,236Z5502G);河北省外专百人计划(360-0803-JSN-3YGS);河北省重点研究开发计划项目(18275501D);河北省教育厅科研项目(QN2022073)。

摘　　要：将跨越式滚环等温扩增技术(SRCA)与荧光技术相结合,建立一种实时荧光SRCA技术检测蜂蜜掺伪大米糖浆的方法。对DNA提取方案进行评估,以大米的特异性基因(PDL、SPS、rbcL、GOS9)为靶序列设计引物,筛选适宜的引物,优化扩增反应条件结合荧光技术建立检测蜂蜜掺伪大米糖浆的方法,并对该方法进行评价。结果表明,建立的实时荧光SRCA方法检测大米DNA的灵敏度为8.45×10~1fg/μL,经特异性评价证实其特异性良好,在人工模拟掺伪检测中建立掺伪比例的对数与Ct值的线性关系,线性方程为y=6.618x+7.651(R~2=0.993),可准确检出蜂蜜中低至1%的大米糖浆成分。该方法灵敏度高,检出限低,能够快速、准确检测蜂蜜掺伪大米糖浆,为蜂蜜掺伪的快速检测提供了新思路。In this study,It combined saltatory rolling circle amplification(SRCA)with fluorescence technology to establish a real-time fluorescence SRCA technique for detection of honey adulterated rice syrup.Three DNA extraction protocols were evaluated,four specific genes of rice(PDL,SPS,rbcL,GOS9)were used as target sequences to design primers,suitable primers were screened,amplification reaction conditions were optimized,and a method for detection of honey adulterated rice syrup was established in combination with fluorescence technology.The results showed that the sensitivity of the established real-time fluorescence SRCA method for the detection of rice DNA was 8.45×101 fg/μL,and its good specificity was confirmed by the specificity evaluation.The linear relationship between the logarithm of the proportion of adulteration and the Ct value was established in the manual simulated adulteration detection test,and the linear equation was y=6.618x+7.651(R2=0.993),which can accurately detect the components of rice syrup in honey down to 1%.The method is sensitive,with low detection limit,and can quickly and accurately quantify the adulteration of honey with rice syrup,which provides a new idea for the rapid quantitative detection of honey adulteration.

关 键 词：蜂蜜掺伪 深加工食品 跨越式滚环等温扩增(SRCA) 

分 类 号：O657.3[理学—分析化学] S896.1[理学—化学]