ADVANCED FUNCTIONAL MATERIALS
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2023
·Vol. 33
·期 7
·2210860
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IF 19
Chenya Zhuo,Huiming Kong,Ke Yi,Chun-Wei Chi,Jiabing Zhang,Ran Chen,Haixia Wang,Caixia Wu,Yeh-Hsing Lao,Yu Tao,Mingqiang Li
Chronic hepatitis B infection remains incurable due to the stable presence of various forms of hepatitis B virus (HBV) genome, especially the HBV covalently closed circular DNA (cccDNA). The emergence of clustered regularly interspaced short palindromic repeat (CRISPR) technology provides a new opportunity to potentially cure the HBV infection. However, the efficiency and specificity remain unsatisfactory, especially for nonviral CRISPR/Cas9 delivery. To tackle these, a liver-specific CRISPR/Cas9 magnetic nanosystem FMNP pAG333/sgXPP is constructed based on fluorinated polyethylenimine-coated magnetic nanoparticles and liver-specific ApoE.HCR.hAAT promoter-driven Cas9-T2A-EGFP plasmid with dual sgRNAs. The elaborate system enables magnetic field-induced spatially specific distribution and hepatocyte-specific promoter-driven liver-specific gene editing. Moreover, this CRISPR/Cas9 magnetic nanosystem is designed to disrupt the two conserved sites in X opening reading frame and Pol opening reading frame of the HBV genome, thereby significantly inactivating the HBV genome without showing off-target effects. Treatment with FMNP pAG333/sgXPP for 7 days reduces serum HBsAg levels by 76% with a total editing efficiency of ≈20% in the two conserved sites. Collectively, this study demonstrates spatiotemporal liver genome editing as well as the feasibility of applying a nonviral CRISPR/Cas9 vector for HBV treatment, which may open up a new application for CRISPR therapeutics.
研究领域:
分子生物学
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纳米技术
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病毒学
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治疗学
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产品关联
DOI
高影响因子
ANALYTICAL CHEMISTRY
·
2023
·
IF 7.4
Zhirun Ji,Bin Zhou,Zhaoyang Shang,Sirui Liu,Xue Li,Xing Zhang,Bingzhi Li
CRISPR-Cas12a is an accurate and responsive biosensing technique, but its limited stability has restricted its widespread applications. To address this, we propose a strategy using metal–organic frameworks (MOFs) to protect Cas12a from harsh environments. After screening multiple candidate MOFs, it was found that hydrophilic MAF-7 is highly compatible with Cas12a, and the as-formed Cas12a-on-MAF-7 (COM) not only retains high enzymatic activity but also possesses excellent tolerance to heat, salt, and organic solvents. Further investigation showed that COM can serve as an analytical component for nucleic acid detection, resulting in an ultrasensitive assay for SARS-CoV-2 RNA detection with a detection limit of 1 copy. This is the first successful attempt to create an active Cas12a nanobiocomposite that functions as a biosensor without the need for shell deconstruction or enzyme release.
研究领域:
生物传感
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纳米技术
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分子生物学
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分析化学
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产品关联
DOI
PubMed
高影响因子
JOURNAL OF HAZARDOUS MATERIALS
·
2021
·
IF 10.588
Pian Wu,Xiaosheng Ye,Danqi Wang,Fangjie Gong,Xiaoqian Wei,Shan Xiang,Jingwen Zhang,Tianhan Kai,Ping Ding
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 (ssDNA-FAM), which was pre-absorbed on Cu-TCPP(Fe) nanosheets. Because of the desorption of the cleaved ssDNA-FAM, 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 μg/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.
研究领域:
分析化学
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生物技术
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环境科学
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DOI
PubMed
高影响因子
INFECTION GENETICS AND EVOLUTION
·
2015
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IF 3.015
Qinqin Huang,Hongping Luo,Minqiang Liu,Jie Zeng,Abualgasim Elgaili Abdalla,Xiangke Duan,Qiming Li,Jianping Xie
Clustered regularly interspaced short palindromic repeats (CRISPR) are present in the genome of 40% bacteria and 90% archaea . CRISPR and accompanying Cas proteins constitute an adaptive immune system against disruptive mobile genetic elements . Two CRISPRs and 9 genes encoding CRISPR-associated proteins have been found in the genome of Mycobacterium tuberculosis . The CRISPR-associated Cas2 is an endoribonuclease required for the acquisition of new spacers. In this study, Cas2 encoded by Rv2816c was expressed in Mycobacterium smegmatis lacking CRISPR-Cas system and its role in stress responses of M. smegmatis in vitro and within macrophages was studied. We found that Cas2 mediated M. smegmatis stress response changes were associated with the altered expression of sigma factors which involved in mycobacterial stress response and virulence. We also found that Cas2 decreased the survival of M. smegmatis within macrophages. This study provides new insights on the role of Cas2.
研究领域:
微生物学
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免疫学
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分子生物学
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DOI
高影响因子
Synthetic and Systems Biotechnology
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2025
·
IF 4.4
Wen Tian,Songcheng Yu,Kaiyang Zhang,Tao Liu,Lihua Ding,Peng Zhang
The CRISPR/Cas12a system holds significant promise for biomedical applications. Nevertheless, the commonly used reporter, fluorophore–quencher-labeled substrates, is hindered by labor-intensive synthesis procedures and high costs, while also relying on a single-photon method and being vulnerable to environmental interference. Herein, a label-free dual-fluorescent functional nucleic acid (DFFNA) was engineered, comprising an aptamer domain for auramine O (AO) recognition and a dSpacer-integrated DNA duplex region for 5,6,7-trimethyl-1,8-naphthyridin-2-amine (ATMND) binding. The fluorescence of AO and ATMND can be enhanced and quenched, respectively, when bound to DFFNAs. The fluorescence intensity ratio between ATMND and AO increased significantly following the cleavage of DFFNAs by activated Cas12a, thus offering a universal, label-free, ratiometric fluorescent reporter for the CRISPR/Cas12a system. To explore the application of the DFFNA-based CRISPR/Cas12a system, a novel biosensor was developed to detect site-specific DNA methylation. It employs a methylation-sensitive restriction enzyme to recognize methylation sites, Cas12a for site-specific DNA identification and signal amplification, and DFFNAs to produce ratiometric fluorescence. The assay demonstrated remarkable specificity and sensitivity, with a limit of detection of 152 pM, due to the high resolution and trans -cleavage activity of Cas12a. The rationally designed and label-free DFFNAs enhance stability, increase flexibility, and reduce cost. The observable color change and smartphone imaging capability facilitate portable, point-of-care testing. Specifically, the biosensor demonstrated excellent specificity by differentiating colorectal cancer patients from healthy individuals. Consequently, this work presents a superior label-free and ratiometric fluorescent reporter for the CRISPR/Cas12a system, which offers a promising strategy for DNA methylation detection in clinical settings.
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DOI
PubMed
高影响因子
ACS Sensors
·
2022
·
IF 9.618
Jiahao Li,Lina Tang,Tingxian Li,Kun Li,Yulin Zhang,Wei Ni,Meng-Meng Xiao,Youyun Zhao,Zhi-Yong Zhang,Guo-Jun Zhang
The path toward field-effect transistor (FET) application from laboratory to clinic has delivered a compelling push in the biomedical domain, yet ultrasensitive and timely pathogen identification without PCR remains a long-lasting challenge. Herein, we create a generic check station termed “CRISPR-FET”, first incorporating the CRISPR/Cas13a system within the FET modality, for accelerated and unamplified detection of viral RNA. Unlike conventional FETs bearing target-specific receptors, this sensor holds three unique advancements: (i) an ingenious sensing mechanism is used, which converts the signal of a large-sized analyte into an on-chip cleavage response of an immobilized CRISPR reporter, enabling signal generation events to occur all within the Debye length; (ii) the multipurpose inspection of the CoV ORF1ab, CoV N gene, and HCV RNA unveils the potential for “one-for-all” scalable FET-based molecular diagnostics; and (iii) it is shown that Cas13a-crRNAs targeting different sites of the viral genome can be deployed in tandem to amplify the FET response, empowering the detection limit down to 1.56 aM, which is a world-record level of sensitivity in the FET for direct viral gene sensing. Notably, a brilliant clinical applicability was made in distinguishing HCV-infected patients from normal controls. Overall, this study sheds new insights into FET-based nucleic acid sensing technology and invokes a vision for its possible future roles in diagnosis of various viral diseases.
研究领域:
生物医学工程
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分子诊断
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纳米技术
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病毒学
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产品关联
DOI
PubMed
高影响因子
Acta Pharmaceutica Sinica B
·
2023
·
IF 14.5
Yongchun Pan,Xiaowei Luan,Fei Zeng,Xuyuan Wang,Shurong Qin,Qianglan Lu,Guanzhong He,Yanfeng Gao,Xiaolian Sun,Xin Han,Bangshun He,Yujun Song
Recent innovations in nanomaterials inspire abundant novel tumor-targeting CRISPR-based gene therapies. However, the therapeutic efficiency of traditional targeted nanotherapeutic strategies is limited by that the biomarkers vary in a spatiotemporal-dependent manner with tumor progression. Here, we propose a self-amplifying logic-gated gene editing strategy for gene/H 2 O 2 -mediated/starvation multimodal cancer therapy. In this approach, a hypoxia-degradable covalent-organic framework (COF) is synthesized to coat a-ZIF-8 in which glucose oxidase (GOx) and CRISPR system are packaged. To intensify intracellular redox dyshomeostasis, DNAzymes which can cleave catalase mRNA are loaded as well. When the nanosystem gets into the tumor, the weakly acidic and hypoxic microenvironment degrades the ZIF-8@COF to activate GOx, which amplifies intracellular H + and hypoxia, accelerating the nanocarrier degradation to guarantee available CRISPR plasmid and GOx release in target cells. These tandem reactions deplete glucose and oxygen, leading to logic-gated-triggered gene editing as well as synergistic gene/H 2 O 2 -mediated/starvation therapy. Overall, this approach highlights the biocomputing-based CRISPR delivery and underscores the great potential of precise cancer therapy.
研究领域:
纳米技术
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癌症生物学
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基因治疗
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生物医学工程
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产品关联
DOI
PubMed
高影响因子
SPECTROCHIMICA ACTA PART A-MOLECULAR AND BIOMOLECULAR SPECTROSCOPY
·
2025
·
IF 4.6
Jiaxuan Chen,Kai Shi
Accurate detection of glyphosate (GLY) residues is essential for protecting public health, ensuring food safety, and preventing environmental pollution caused by excessive or improper herbicide application. Based on the asymmetric CRISPR/Cas12a system as a signal amplification technique, with phosphorothioate-modified hairpin G-quadruplex (psHG4) acting as a signal probe, a platform named ACC-GLY is developed for the sensitive and accurate detection of GLY. In the designed ACC-GLY platform, the target GLY specifically binds to the aptamer, and the cascading signal amplification strategy, driven by the DNase activity of a single Cas12a, is initiated by two competitive guide RNAs. Under the influence of Cas12a's DNase activity, the psHG4 probe is cleaved, releasing the psG4 sequence. The released psG4 sequence then binds to Thioflavin T (ThT), forming a complex that generates a fluorescence signal. Under optimal conditions, the detection platform can specifically detect GLY at a concentration as low as 0.3 pM. The detection platform demonstrates significant capability in detecting GLY in tap water and corn samples, highlighting its vast potential for applications in environmental monitoring and food safety. To the best of our knowledge, this is the first study to utilize the CRISPR/Cas12a system for GLY detection.
研究领域:
分析化学
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生物传感器
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环境监测
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食品安全
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分子生物学
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产品关联
DOI
PubMed
高影响因子
Food Frontiers
·
2023
·
IF 9.9
Yaru Li,Jiali Qiao,Zhiying Zhao,Qiang Zhang,Wenlu Zhang,Shuli Man,Shengying Ye,Kai Chen,Long Ma
It is imperative to develop practicable pathogenic bacteria detection methods. We devised a biosensor for the ultrasensitive detection of Salmonella typhimurium ( S. typhi ), termed as SCOUT-dCas9 (ultrasensitive, cross-validating, on-site, and dUal-mode test using CRISPR/dCas9). Simply, the species-specific invA gene of S. typhi was amplified using loop-mediated isothermal amplification with a biotinylated primer, which can be specifically “pulled down” by “antibody-like” dCas9-single guide RNA to form a ternary complex. SYBR Green I and streptavidin-modified alkaline phosphatase were used to functionalize them to generate fluorescent and colorimetric signals, respectively. With this strategy, the target could be dexterously converted into bimodal signals that were cross-validated to afford more reliable results. For both modes, SCOUT-dCas9 was able to detect as low as 1 CFU/mL with a dynamic range from 1 to 10 9 CFU/mL. Lastly, SCOUT-dCas9 had satisfactory selectivity and was capable of detecting S. typhi -contaminated real food samples. SCOUT-dCas9 provides a robust platform for bacterial detection.
研究领域:
生物技术
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微生物学
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食品科学
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DOI
高影响因子
TALANTA
·
2025
·
IF 6.1
Haixiang Li,Shuo Lian,Zhiwei Zhang,Weiye Bi,Qingyou Meng,Qingwei Ding
Brain natriuretic peptide (BNP) is considered a reliable marker of heart failure disease, and its timely detection can provide important pathological information to prevent or treat heart failure. In this article, an electrochemiluminescence (ECL) sensing device based on a boron carbon nitride/gold nanoparticle (BCN/AuNPs) complex is developed to determine BNP. Prominently, the CRISPR/CAS 13a enzyme reverse cleavage mode, the entropy-driven and hammer hybridization chain reaction processes were involved in the entire detection scheme. Ultimately, with multiple reaction methods and amplification reactions of nucleic acids, this ECL sensing device is able to achieve a detection limit as low as of 0.03 pg/mL and linear range from 0.1 pg/mL to 30 ng/mL for BNP. In addition, the ECL sensing device based on BCN/AuNPs complex obtained satisfactory stability and specificity, and can also be extended to the detection of other pathological markers.
研究领域:
生物医学工程
·
分子诊断学
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产品关联
DOI
PubMed
高影响因子
ACS Nano
·
2024
·
IF 15.8
Xiaoqi Tang,Yihui Chen,Binpan Wang,Dan Luo,Jue Wang,Yuan He,Liu Feng,Ying Xu,Shuang Xie,Ming Chen,Kai Chang
Manipulating the expression of cellular genes through efficient CRISPR/Cas9 delivery is rapidly evolving into a desirable tumor therapeutics. The exposure of CRISPR/Cas9 to a complex external environment poses challenges for conventional delivery carriers in achieving responsive and accurate release. Here, we report a Trojan horse-like nanocapsule for the on-demand delivery of CRISPR/Cas9 in a microRNA-responsive manner, enabling precise tumor therapy. The nanocapsule comprises a nanoassembled, engineered DNAzyme shell encasing a Cas9/sgRNA complex core. The DNAzyme, functioning as a catalytic unit, undergoes a conformational change in the presence of tumor-associated microRNA, followed by activating a positive feedback-driven autonomous catabolic cycle of the nanocapsule shell. This catabolic cycle is accomplished through chain reactions of DNAzyme “cleavage–hybridization–cleavage”, which ensures sensitivity in microRNA recognition and effective release of Cas9/sgRNA. Utilizing this Trojan horse-like nanocapsule, as low as 1.7 pM microRNA-21 can trigger the on-demand release of Cas9/sgRNA, enabling the specific editing of the protumorigenic microRNA coding gene. The resulting upregulation of tumor suppressor genes induces apoptosis in tumor cells, leading to significant inhibition of tumor growth by up to 75.94%. The Trojan horse-like nanocapsule, with superior programmability and biocompatibility, is anticipated to serve as a promising carrier for tailoring responsive gene editing systems, achieving enhanced antitumor specificity and efficacy.
研究领域:
生物技术
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纳米技术
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癌症研究
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基因治疗
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分子生物学
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产品关联
DOI
PubMed
高影响因子
Advanced Science
·
2020
·Vol. 7
·期 14
·1903381
·
IF 15.84
Hang Yin,Xiaoling Yuan,Lihua Luo,Yichao Lu,Bing Qin,Junlei Zhang,Yingying Shi,Chunqi Zhu,Jie Yang,Xiang Li,Mengshi Jiang,Zhenyu Luo,Xinyu Shan,Dawei Chen,Jian You
The development of gene delivery has attracted increasing attention, especially when the introduction and application of the CRISPR/Cas9 gene editing system appears promising for gene therapy. However, ensuring biosafety and high gene editing efficiency at the same time poses a great challenge for its in vivo applications. Herein, a pardaxin peptide (PAR)-modified cationic liposome (PAR-Lipo) is developed. The results are indicative that significantly enhanced gene editing efficiency can be obtained through the mediation of PAR-Lipos compared to non-Lipos (non-PAR-modified liposomes) and Lipofectamine 2000, owing to its protection toward carried nucleotide by the prevention of lysosomal capture, prolongation of retention time in cells through the accumulation in the endoplasmic reticulum (ER), and more importantly, facilitation of the nuclear access via an ER-nucleus route. Accumulation of PAR-Lipos in the ER may improve the binding of Cas9 and sgRNA, thus further contributing to the eventually enhanced gene editing efficiency. Given their high biosafety, PAR-Lipos are used to mediate the knockout of the oncogene CDC6 in vivo, which results in significant tumor growth inhibition. This work may provide a useful reference for enhancing the delivery of gene editing systems, thus improving the potential for their future clinical applications.
研究领域:
生物技术
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基因治疗
·
分子生物学
·
癌症研究
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产品关联
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）