共 50 条
Portable biosensor combining CRISPR/Cas12a and loop-mediated isothermal amplification for antibiotic resistance gene ermB in wastewater
被引:21
作者:
Mao, Kang
[1
]
Zhang, Hua
[1
]
Ran, Fang
[1
]
Cao, Haorui
[1
]
Feng, Rida
[1
]
Du, Wei
[2
]
Li, Xiqing
[3
]
Yang, Zhugen
[4
]
机构:
[1] Chinese Acad Sci, Inst Geochem, State Key Lab Environm Geochem, Guiyang 550081, Peoples R China
[2] Kunming Univ Sci & Technol, Fac Environm Sci & Engn, Kunming 650500, Peoples R China
[3] Peking Univ, Coll Urban & Environm Sci, Lab Earth Surface Proc, Beijing 100871, Peoples R China
[4] Cranfield Univ, Sch Water Energy & Environm, Cranfield MK43 0AL, England
基金:
中国国家自然科学基金;
关键词:
Antibiotic resistance gene;
CRISPR/Cas12a;
Loop-mediated isothermal amplification;
Biosensor;
Wastewater;
NUCLEIC-ACID DETECTION;
DIAGNOSTICS;
SARS-COV-2;
D O I:
10.1016/j.jhazmat.2023.132793
中图分类号:
X [环境科学、安全科学];
学科分类号:
08 ;
0830 ;
摘要:
Wastewater is among the main sources of antibiotic resistance genes (ARGs) in the environment, but effective methods to quickly assess ARGs on-site in wastewater are lacking. Here, using the typical ARG ermB as the target, we report a portable biosensor combining CRISPR/Cas12a and loop-mediated isothermal amplification (LAMP) for the detection of ARGs. Six primers of LAMP and the crRNA of CRISPR/Cas12a were first designed to be preamplification with LAMP and lead Cas12a to recognize the ermB via base pairing. Due to the trans-cleavage activity of CRISPR/Cas12a after amplicon recognition, ssDNA probes modified with reporter molecules were used to implement a visual assay with lateral flow test strips and fluorescence. After a simple nucleic acid extraction with magnetic beads, the constructed biosensor possesses excellent sensitivity and selectivity as low as 2.75 x 103 copies/mu L using fluorescence and later flow strips in wastewater. We further evaluated the community-wide prevalence of ermB in wastewater influent and found high mass loads of ermB during different months. This user-friendly and low-cost biosensor is applicable for rapid on-site ARG detection, providing a
引用
收藏
页数:10
相关论文
共 50 条