Rapid and multiplex preparation of engineered <i>Mycobacterium smegmatis</i> porin A (MspA) nanopores for single molecule sensing and sequencing

Yan, Shuanghong; Wang, Liying; Du, Xiaoyu; Zhang, Shanyu; Wang, Sha; Cao, Jian; Zhang, Jinyue; Jia, Wendong; Wang, Yuqin; Zhang, Panke; Chen, Hong‐Yuan; Huang, Shuo

doi:10.1039/d1sc01399h

Cited by 9 publications

(3 citation statements)

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“…Moreover, the MspA nanopore can distinguish all four nucleobases and achieve a significantly more difference in ionic current between nucleobases than α-hemolysin ( Laszlo et al, 2016 ). Further research has developed new insights into making a variety of MspA mutants that can rapidly and inexpensively produce MspA protein nanopores, and the performance of nanopores prepared by this method is not different from that of nanopores prepared by other methods ( Yan S. et al, 2021 );…”

Section: The History Of Biological Nanopore Sequencingmentioning

confidence: 99%

Portable nanopore-sequencing technology: Trends in development and applications

et al. 2023

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Sequencing technology is the most commonly used technology in molecular biology research and an essential pillar for the development and applications of molecular biology. Since 1977, when the first generation of sequencing technology opened the door to interpreting the genetic code, sequencing technology has been developing for three generations. It has applications in all aspects of life and scientific research, such as disease diagnosis, drug target discovery, pathological research, species protection, and SARS-CoV-2 detection. However, the first- and second-generation sequencing technology relied on fluorescence detection systems and DNA polymerization enzyme systems, which increased the cost of sequencing technology and limited its scope of applications. The third-generation sequencing technology performs PCR-free and single-molecule sequencing, but it still depends on the fluorescence detection device. To break through these limitations, researchers have made arduous efforts to develop a new advanced portable sequencing technology represented by nanopore sequencing. Nanopore technology has the advantages of small size and convenient portability, independent of biochemical reagents, and direct reading using physical methods. This paper reviews the research and development process of nanopore sequencing technology (NST) from the laboratory to commercially viable tools; discusses the main types of nanopore sequencing technologies and their various applications in solving a wide range of real-world problems. In addition, the paper collates the analysis tools necessary for performing different processing tasks in nanopore sequencing. Finally, we highlight the challenges of NST and its future research and application directions.

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Section: The History Of Biological Nanopore Sequencingmentioning

confidence: 99%

Portable nanopore-sequencing technology: Trends in development and applications

et al. 2023

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“…The characteristic event features acquired at the beginning of the measurements and a few hours later also appear completely identical (Supplementary Figs 38, 39). Though the MspA nanopore is protein in nature, according to previous investigations [58][59][60] , it demonstrates a high rigidity and durability in the structure, which is ideal for long term measurement and device storage.…”

Section: Rapid Analysis Of Natural Fruit Juice By a Nanoporementioning

confidence: 99%

Nanopore analysis of cis-diols in fruits

Fan,

Cao,

Zhang

et al. 2024

Nat Commun

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Natural fruits contain a large variety of cis-diols. However, due to the lack of a high-resolution sensor that can simultaneously identify all cis-diols without a need of complex sample pretreatment, direct and rapid analysis of fruits in a hand-held device has never been previously reported. Nanopore, a versatile single molecule sensor, can be specially engineered to perform this task. A hetero-octameric Mycobacterium smegmatis porin A (MspA) nanopore modified with a sole phenylboronic acid (PBA) adapter is prepared. This engineered MspA accurately recognizes 1,2-diphenols, alditols, α-hydroxy acids and saccharides in prune, grape, lemon, different varieties of kiwifruits and commercial juice products. Assisted with a custom machine learning program, an accuracy of 99.3% is reported and the sample pretreatment is significantly simplified. Enantiomers such as DL-malic acids can also be directly identified, enabling sensing of synthetic food additives. Though demonstrated with fruits, these results suggest wide applications of nanopore in food and drug administration uses.

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“…Because the amino acid residues around the pore constriction zone significantly affect sensing accuracy, various mutants have been obtained for different target molecules. For instance, M2 MspA (D90 N/D91 N/D93 N/D118R/E139 K/D134R) has been applied to detect longer peptide chains; and MspA−C, MspA−H, MspA−M, and MspA−D have mutated the narrowest restriction‐1.2 nm wide of MspA at the position 91 to different amino acids to study the coordination interactions between amino acids and metal ions [41] . Overall, MspA mutants have been investigated for DNA sequencing, [4b] epigenetic sequencing, [42] miRNA sequencing, [43] DNA lesion sequencing, [44] and protein sequencing [45] …”

Section: Nanoporementioning

confidence: 99%

Recent Advances in Aptamer‐Based Nanopore Sensing at Single‐Molecule Resolution

Zhang

Yang

et al. 2022

Chemistry An Asian Journal

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Aptamers as the recognition element of the stochastic nanopore sensors have been under intense investigation. This paper reviews recent research advances in aptamer‐based nanopore sensing techniques, including the classification and selection of nanopores (biological nanopores, solid‐state nanopores, and nanopipettes), different strategies of aptamer‐based nanopore sensing, and their values and outlook for applications in areas such as environmental analysis, precision diagnosis, pharmaceutical industry, and security. Furthermore, the single‐molecule nanopore sensors have been applied to reveal the aptamer‐target interactions, such as recognition orientation, binding sites, and conformational heterogeneity dissociation kinetics at the single‐molecule level. In this review, recent research efforts to develop aptamer‐based single‐molecule nanopore sensors with high selectivity and sensitivity are highlighted, and some perspectives are drawn.

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Rapid and multiplex preparation of engineered Mycobacterium smegmatis porin A (MspA) nanopores for single molecule sensing and sequencing

Abstract: Acknowledging its unique conical lumen structure, Mycobacterium smegmatis porin A (MspA) was the first type of nanopore that has successfully sequenced DNA. Recent developments of nanopore single molecule chemistry have...

Cited by 9 publications

References 29 publications

Portable nanopore-sequencing technology: Trends in development and applications

Portable nanopore-sequencing technology: Trends in development and applications

Nanopore analysis of cis-diols in fruits

Recent Advances in Aptamer‐Based Nanopore Sensing at Single‐Molecule Resolution

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