A novel CRISPR/Cas14a system integrated with 2D porphyrin metal-organic framework for microcystin-LR determination through a homogeneous competitive reaction
A novel CRISPR/Cas14a system integrated with 2D porphyrin metal-organic framework for microcystin-LR determination through a homogeneous competitive reaction
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一种与 2D 卟啉金属有机框架集成的新型 CRISPR/Cas14a 系统，用于通过均相竞争反应测定微囊藻毒素-LR
DOI：
10.1016/j.jhazmat.2021.127690
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发表时间：
2021-12-09
影响因子：
13.6
通讯作者：
Ding, Ping
中科院分区：
文献类型：
--
作者：
Wu, Pian;Ye, Xiaosheng;Ding, Ping
微囊藻毒素-LR(MC-LR)毒性大、分布广，对其进行选择性、灵敏的检测具有重要意义。本文将CRISPR/Cas14a体系与二维金属有机骨架纳米片(2D-PMOFS)相结合，开发了一种新型的多功能荧光传感器(Cas14-PMOFS荧光传感器)，用于MC-LR的测定。设计的CRISPR/Cas14a系统可被未结合的互补DNA激活，并与MC-LR浓度呈正相关。此外，激活的Cas14a蛋白被用来不分青红皂白地切割预先吸附在Cu-TCPP(Fe)纳米片上的FAM标记的单链DNA(SsDNAFAM)。由于裂解的ssDNAFAM解吸，恢复了预猝灭的荧光信号。由于这种Cas14-pMOFS荧光传感器对c DNA具有良好的定量性能，其检测下限(LOD)为0.12 nM，因此该荧光传感器能够在50 pg/m L到1 mU g/m L范围内检测MC-LR，检测下限为19 pg/m L。这项工作不仅为基于2D-PMOF与CRISPR/Cas14a耦合的荧光传感器的探索提供了新的视角，而且展示了其在核酸和非核酸靶标检测中的普适性。
Selective and sensitive detection of microcystin-LR (MC-LR) is of vital importance because of its high toxicity and broad distribution. Herein, a novel and versatile fluorescence sensor (Cas14-pMOFs fluorescence sensor) was developed by combining the CRISPR/Cas14a system with a 2D porphyrin metal-organic framework nanosheets (2D-pMOFs) for MC-LR determination. The designed CRISPR/Cas14a system was activated by the unbound complementary DNA (cDNA), which was positively correlated with MC-LR concentration. Furthermore, the activated Cas14a protein was utilized to indiscriminately cleave the FAM-labeled single-stranded DNA (ssDNAFAM), which was pre-absorbed on Cu-TCPP(Fe) nanosheets. Because of the desorption of the cleaved ssDNAFAM, the pre-quenched fluorescence signal was recovered. Owing to the excellent performance in quantifying cDNA using this Cas14-pMOFs fluorescence sensor with a limit of detection (LOD) of 0.12 nM, this Cas14-pMOFs fluorescence sensor was able to detect MC-LR in a range from 50 pg/mL to 1 mu g/mL with the LOD of 19 pg/mL. This work not only provided a new insight for the exploration of fluorescence sensors based on 2D-pMOFs coupled with CRISPR/Cas14a, but also, demonstrated its universality in both nucleic acid and non-nucleic acid targets determination.