基于核酸适配体传感器检测食品致病菌的研究进展

doi:10.13560/j.cnki.biotech.bull.1985.2019-1273

摘要/Abstract

摘要：

核酸适配体是可以识别并与靶标分子结合的单链DNA或RNA分子。由于适配体具有良好的特异性、稳定性、选择性、易于修饰和亲和力高等特点,在病原检测和其他小分子识别方面具有很大的潜力。综述了适配体的筛选技术以及适配体应用技术,包括比色法、可见光谱法、荧光法、电化学法、表面增强拉曼法和胶体金试纸条法等在食源性致病菌检测中的应用。目前适配体传感器在检测食品致病菌领域取得了显著进展。然而核酸适配体应用方面的挑战和缺陷仍需克服。

关键词: 适配体, 食品安全, 快速检测, SELEX, 大肠杆菌, 沙门氏菌

Abstract:

Aptamers are single-stranded DNA or RNA molecules which possess specific identification and binding ability to their targets. Aptamers present breathtaking potential in the field of foodborne pathogen detection and other bio-molecular recognition,owing to their excellent specificity,stabilization,selectivity,simplicity of modification,and high affinity. This review outlines the technologies of aptamer screening and aptamer application,including colorimetric assays,visible spectrum assay,fluorescence assays,electrochemical assays,surface-enhanced Raman assays,and gold-nanoparticle lateral flow assays in the determination of foodborne pathogens. Until now,the remarkable progress（more rapid,sensitive,and accurate）in the monitoring of foodborne pathogen on aptasensor has been achieved;however,there are still many challenges and flaws in their utilization to be overcome.

Key words: aptamer, food safety, rapid detection, SELEX, Escherichia coli, Salmonella

王琦, 颜春蕾, 高洪伟, 吴薇, 杨庆利. 基于核酸适配体传感器检测食品致病菌的研究进展[J]. 生物技术通报, 2020, 36(11): 245-258.

WANG Qi, YAN Chun-lei, GAO Hong-wei, WU Wei, YANG Qing-li. Research Progress of DNA Aptasensors for Foodborne Pathogen Detection[J]. Biotechnology Bulletin, 2020, 36(11): 245-258.

图/表 9

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靶标	菌型	筛选方法	序列（5'-3'）	参考文献
大肠杆菌	大肠杆菌O157：H7	全细胞-SELEX	ACCAGTAGACTTTCAACTTTACTGCCATCGTGTGCCCTAA	[25,32]
	大肠杆菌O157：H7	Non-SELEX	TATGGCGTGGCAAGCTTGGCCCGCTTCTCAAGCATGGTTATCTAC	[33]
	大肠杆菌O111：B4	磁珠-SELEX	ATCCGTCACCCCTGCTCTCGTCGCTATGAAGTAACAAAGATAGGAGCAAT CGGGTGGTGTTGGCTCCCGTAT	[34]
	大肠杆菌O111	全细胞-SELEX	ATCCGTCACACCTGCTCTACTGGCCGGCTCAGCATGACTAAGAAGGAAGT TATGTGGTGTTGGCTCCCGTAT	[35]
	大肠杆菌O55：B5	Non-SELEX	TAGCCGGATCGCGCTGGCCAGATGATATAAAGGGTCAGCCCCCCA	[36]
沙门氏菌	鼠伤寒沙门氏菌	全细胞-SELEX	ACGGGCGTGGGGGCAATGCCTGCTTGTAGGCTTCCCCTGTGCGCG	[37]
	鼠伤寒沙门氏菌	SELEX	TATGGCGGCGTCACCCGACGGGGACTTGACATTATGACAG	[38]
	沙门氏菌O8	全细胞-SELEX	GATCCGGGCCTCATGTCGAACACCCCCCAACTAAAACAACAAAACACC ACCGCCATTGAGCGTTTATTCTGAGCTCCCA	[39]
	沙门氏菌O8	SELEX	TGATCCGGGCCTCATGTCGAACACCCCCCAACTAAAACAACAAAACAC CACCGCCATTGAGCGTTTATTCTGGCTCCCA	[40]
	肠炎沙门氏菌	磁珠-SELEX	GGGUUCACUGCAGACUUGACGAAGCUUGAGAGAUGCCCCCUGAUGTGC AUUCUUGUUGUGUUGCGGCAAUGGAUCCACAUCTACGAAUUC	[41]
金黄色葡萄球菌	金黄色葡萄球菌	全细胞-SELEX	GCGCCCTCTCACGTGGCACTCAGAGTGCCGGAAGTTCTGCGTTAT	[42]
	肠毒素	SELEX	TTTGGTATTGAGGGTCGCATCCACTGGTCGTTGTCTGTTGTCTGTTAT GTTGTTTCGTGATGGCTCTAACTCTCCTCT	[43]
	磷壁酸	SELEX	GGGAGUUUUGAUACGGCUUCAUGCAGUAAUGUUUUUAU	[31]
	肠毒素B	SELEX	AGCAGCACAGAGGTCAGATGTATACTTCTAAAATTTGTTTGTATCTAC GATGTTCTTCGTCCTATGCGTGCTACCGTGAA	[44]
	肠毒素A	SELEX	TACTTATGCATTTCCTCCCACGATCTTATTTGAGAGTGAC	[45]
副溶血弧菌		全细胞-SELEX	ATAGGAGTCACGACGACCAGAATCTAAAAATGGGCAAAGAAACAGT GACTCGTTGAGATACTTATGTGCGTCTACCTCTTGACTAAT	[46]
单核细胞增生李斯特菌		全细胞-SELEX	TGGGAGCTCAGAATAAACGCTCAACTTTGTTCTTCTTTGCTTTTTTTTT CTTTTTTTGTTCGACATGAGGCCCGGATCA	[47]
蜡样芽孢杆菌	芽孢	SELEX	CATCCGTCACACCTGCTCGGTGCAGACCCATAGGGGGGGCGTGCGGA TGTAGGAGTAGGGTGTTGGCTCCCGTATC	[48]
志贺氏菌	痢疾志贺氏菌	全细胞-SELEX	CGGAACTAGCGTTTAAATGCCAGGACTGAAGTAGGCAGGG	[22]
志贺氏菌	宋内志贺菌	全细胞-SELEX	ATTATGTTGTCTGAACTGCCATGGTCCCTCGTGTTTATTATGTTGTCT GACTGGCTGAGATTGCACTTACTATCT	[49]
绿脓杆菌		全细胞-SELEX	CCCCCGTTGCTTTCGCTTTTCCTTTCGCTTTTGTTCGTTTCGTCCCTGC TTCCTTTCTTG	[50]

靶标	菌型	筛选方法	序列（5'-3'）	参考文献
大肠杆菌	大肠杆菌O157：H7	全细胞-SELEX	ACCAGTAGACTTTCAACTTTACTGCCATCGTGTGCCCTAA	[25,32]
	大肠杆菌O157：H7	Non-SELEX	TATGGCGTGGCAAGCTTGGCCCGCTTCTCAAGCATGGTTATCTAC	[33]
	大肠杆菌O111：B4	磁珠-SELEX	ATCCGTCACCCCTGCTCTCGTCGCTATGAAGTAACAAAGATAGGAGCAAT CGGGTGGTGTTGGCTCCCGTAT	[34]
	大肠杆菌O111	全细胞-SELEX	ATCCGTCACACCTGCTCTACTGGCCGGCTCAGCATGACTAAGAAGGAAGT TATGTGGTGTTGGCTCCCGTAT	[35]
	大肠杆菌O55：B5	Non-SELEX	TAGCCGGATCGCGCTGGCCAGATGATATAAAGGGTCAGCCCCCCA	[36]
沙门氏菌	鼠伤寒沙门氏菌	全细胞-SELEX	ACGGGCGTGGGGGCAATGCCTGCTTGTAGGCTTCCCCTGTGCGCG	[37]
	鼠伤寒沙门氏菌	SELEX	TATGGCGGCGTCACCCGACGGGGACTTGACATTATGACAG	[38]
	沙门氏菌O8	全细胞-SELEX	GATCCGGGCCTCATGTCGAACACCCCCCAACTAAAACAACAAAACACC ACCGCCATTGAGCGTTTATTCTGAGCTCCCA	[39]
	沙门氏菌O8	SELEX	TGATCCGGGCCTCATGTCGAACACCCCCCAACTAAAACAACAAAACAC CACCGCCATTGAGCGTTTATTCTGGCTCCCA	[40]
	肠炎沙门氏菌	磁珠-SELEX	GGGUUCACUGCAGACUUGACGAAGCUUGAGAGAUGCCCCCUGAUGTGC AUUCUUGUUGUGUUGCGGCAAUGGAUCCACAUCTACGAAUUC	[41]
金黄色葡萄球菌	金黄色葡萄球菌	全细胞-SELEX	GCGCCCTCTCACGTGGCACTCAGAGTGCCGGAAGTTCTGCGTTAT	[42]
	肠毒素	SELEX	TTTGGTATTGAGGGTCGCATCCACTGGTCGTTGTCTGTTGTCTGTTAT GTTGTTTCGTGATGGCTCTAACTCTCCTCT	[43]
	磷壁酸	SELEX	GGGAGUUUUGAUACGGCUUCAUGCAGUAAUGUUUUUAU	[31]
	肠毒素B	SELEX	AGCAGCACAGAGGTCAGATGTATACTTCTAAAATTTGTTTGTATCTAC GATGTTCTTCGTCCTATGCGTGCTACCGTGAA	[44]
	肠毒素A	SELEX	TACTTATGCATTTCCTCCCACGATCTTATTTGAGAGTGAC	[45]
副溶血弧菌		全细胞-SELEX	ATAGGAGTCACGACGACCAGAATCTAAAAATGGGCAAAGAAACAGT GACTCGTTGAGATACTTATGTGCGTCTACCTCTTGACTAAT	[46]
单核细胞增生李斯特菌		全细胞-SELEX	TGGGAGCTCAGAATAAACGCTCAACTTTGTTCTTCTTTGCTTTTTTTTT CTTTTTTTGTTCGACATGAGGCCCGGATCA	[47]
蜡样芽孢杆菌	芽孢	SELEX	CATCCGTCACACCTGCTCGGTGCAGACCCATAGGGGGGGCGTGCGGA TGTAGGAGTAGGGTGTTGGCTCCCGTATC	[48]
志贺氏菌	痢疾志贺氏菌	全细胞-SELEX	CGGAACTAGCGTTTAAATGCCAGGACTGAAGTAGGCAGGG	[22]
志贺氏菌	宋内志贺菌	全细胞-SELEX	ATTATGTTGTCTGAACTGCCATGGTCCCTCGTGTTTATTATGTTGTCT GACTGGCTGAGATTGCACTTACTATCT	[49]
绿脓杆菌		全细胞-SELEX	CCCCCGTTGCTTTCGCTTTTCCTTTCGCTTTTGTTCGTTTCGTCCCTGC TTCCTTTCTTG	[50]

传感器	适用场所	灵敏度CFU/mL	优缺点	靶标	LOD	检测范围	参考文献
比色适配体传感器	直观快速检测复杂的食品基质	10	结果直观,适用于现场检测,但结果难以量化	副溶血性弧菌	10 CFU/mL	10²-10⁷CFU/mL	[57]
比色适配体传感器	直观快速检测复杂的食品基质	10	结果直观,适用于现场检测,但结果难以量化	鼠伤寒沙门氏菌	11 CFU/mL	11-1.10×10⁵CFU/mL	[58]
可见光适配体传感器	POC测试与商业检测	100	检测快速、易于操作、成本低、仪器便携,但灵敏度较差,难以精确量化	伤寒沙门菌	10³CFU/mL	10³-10⁷ CFU /mL	[59]
可见光适配体传感器	POC测试与商业检测	100	检测快速、易于操作、成本低、仪器便携,但灵敏度较差,难以精确量化	金黄色葡萄球菌	3.17 μmol/L	8-23 μmol/L	[60]
荧光适配体传感器	各种高通量的样品	10³	特异性高,试剂和仪器成本高	嗜水气单胞菌和迟缓爱德华菌	1.5 CFU/mL和 67 CFU/mL	0.1-1.3×10⁵CFU/mL和1.3-1.3×10⁴CFU/mL	[61]
电化学适配体传感器	痕量检测			金黄色葡萄球菌	1 CFU/mL	10-1×10⁶CFU/mL	[62]
电化学适配体传感器	痕量检测			O157：H7型大肠杆菌	10 CFU/mL	-	[56]
表面增强拉曼散射适配体传感器	痕量靶标的无损检测	5	灵敏度高,无标签,专业性要求高,仪器贵重	金黄色葡萄球菌	1.5 CFU/mL	10-10⁷CFU/mL	[63]
测流适配体传感器	直观快速检测	10	方便携带、易于操作、成本低,但难以量化	副溶血性弧菌	5.6 CFU/mL	-	[19]

传感器	适用场所	灵敏度CFU/mL	优缺点	靶标	LOD	检测范围	参考文献
比色适配体传感器	直观快速检测复杂的食品基质	10	结果直观,适用于现场检测,但结果难以量化	副溶血性弧菌	10 CFU/mL	10²-10⁷CFU/mL	[57]
比色适配体传感器	直观快速检测复杂的食品基质	10	结果直观,适用于现场检测,但结果难以量化	鼠伤寒沙门氏菌	11 CFU/mL	11-1.10×10⁵CFU/mL	[58]
可见光适配体传感器	POC测试与商业检测	100	检测快速、易于操作、成本低、仪器便携,但灵敏度较差,难以精确量化	伤寒沙门菌	10³CFU/mL	10³-10⁷ CFU /mL	[59]
可见光适配体传感器	POC测试与商业检测	100	检测快速、易于操作、成本低、仪器便携,但灵敏度较差,难以精确量化	金黄色葡萄球菌	3.17 μmol/L	8-23 μmol/L	[60]
荧光适配体传感器	各种高通量的样品	10³	特异性高,试剂和仪器成本高	嗜水气单胞菌和迟缓爱德华菌	1.5 CFU/mL和 67 CFU/mL	0.1-1.3×10⁵CFU/mL和1.3-1.3×10⁴CFU/mL	[61]
电化学适配体传感器	痕量检测			金黄色葡萄球菌	1 CFU/mL	10-1×10⁶CFU/mL	[62]
电化学适配体传感器	痕量检测			O157：H7型大肠杆菌	10 CFU/mL	-	[56]
表面增强拉曼散射适配体传感器	痕量靶标的无损检测	5	灵敏度高,无标签,专业性要求高,仪器贵重	金黄色葡萄球菌	1.5 CFU/mL	10-10⁷CFU/mL	[63]
测流适配体传感器	直观快速检测	10	方便携带、易于操作、成本低,但难以量化	副溶血性弧菌	5.6 CFU/mL	-	[19]