Ultrasensitive, rapid, and highly specific detection of microRNAs based on PER-CRISPR/CAS
Abnormal microRNA (miRNA) expression levels are confirmed as diagnostic biomarkers of the emergence and development of diseases. In this study, we developed a fluorescence biosensor for detecting miRNAs based on double amplification reactions with the primer exchange reaction (PER) and CRISPR/Cas12a...
Ausführliche Beschreibung
Autor*in: |
Wang, Ze [verfasserIn] |
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Englisch |
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2022transfer abstract |
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Übergeordnetes Werk: |
Enthalten in: Feeding European sea bass ( - Torrecillas, S. ELSEVIER, 2018, a Tetrahedron publication for rapid dissemination of preliminary communications on all aspects of bioorganic chemistry, medicinal chemistry, bioinorganic chemistry and related disciplines, Amsterdam [u.a.] |
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volume:74 ; year:2022 ; day:15 ; month:10 ; pages:0 |
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DOI / URN: |
10.1016/j.bmcl.2022.128949 |
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ELV058702350 |
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520 | |a Abnormal microRNA (miRNA) expression levels are confirmed as diagnostic biomarkers of the emergence and development of diseases. In this study, we developed a fluorescence biosensor for detecting miRNAs based on double amplification reactions with the primer exchange reaction (PER) and CRISPR/Cas12a. In the absence of target miRNA-21, PER hairpins remained locked by the protector strands and the primers did not extend. In the presence of target miRNA-21, the miRNA-21 bound to the guard sequence and exposed primer binding sites. Also, the closed PER hairpin was unlocked to specifically extend primers into single-stranded DNA (ssDNA) of unequal lengths. These ssDNAs of unequal lengths could activate the cleavage of a reporter by Cas12a, leading to an increase in detectable fluorescence signals. A large number of short nucleic acid fragments were amplified by PER-CRISPR multiple cycle cleavage fluorescent probes. Based on PER-combined CRISPR/Cas12a established dual signal amplification method was characterized by a low limit of detection of 10fM. The fluorescent biosensor for miRNA detection had the advantages of low detection cost, simple operation, and mobility, providing a very promising platform for the point-of-care testing of miRNA-21. | ||
520 | |a Abnormal microRNA (miRNA) expression levels are confirmed as diagnostic biomarkers of the emergence and development of diseases. In this study, we developed a fluorescence biosensor for detecting miRNAs based on double amplification reactions with the primer exchange reaction (PER) and CRISPR/Cas12a. In the absence of target miRNA-21, PER hairpins remained locked by the protector strands and the primers did not extend. In the presence of target miRNA-21, the miRNA-21 bound to the guard sequence and exposed primer binding sites. Also, the closed PER hairpin was unlocked to specifically extend primers into single-stranded DNA (ssDNA) of unequal lengths. These ssDNAs of unequal lengths could activate the cleavage of a reporter by Cas12a, leading to an increase in detectable fluorescence signals. A large number of short nucleic acid fragments were amplified by PER-CRISPR multiple cycle cleavage fluorescent probes. Based on PER-combined CRISPR/Cas12a established dual signal amplification method was characterized by a low limit of detection of 10fM. The fluorescent biosensor for miRNA detection had the advantages of low detection cost, simple operation, and mobility, providing a very promising platform for the point-of-care testing of miRNA-21. | ||
650 | 7 | |a miRNA-21 |2 Elsevier | |
650 | 7 | |a Fluorescence |2 Elsevier | |
650 | 7 | |a PER |2 Elsevier | |
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700 | 1 | |a Wei, Hongguo |4 oth | |
700 | 1 | |a Bu, Shengjun |4 oth | |
700 | 1 | |a Li, Xue |4 oth | |
700 | 1 | |a Zhou, Hongyu |4 oth | |
700 | 1 | |a Zhang, Wenhui |4 oth | |
700 | 1 | |a Wan, Jiayu |4 oth | |
773 | 0 | 8 | |i Enthalten in |n Elsevier Science |a Torrecillas, S. ELSEVIER |t Feeding European sea bass ( |d 2018 |d a Tetrahedron publication for rapid dissemination of preliminary communications on all aspects of bioorganic chemistry, medicinal chemistry, bioinorganic chemistry and related disciplines |g Amsterdam [u.a.] |w (DE-627)ELV000272361 |
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10.1016/j.bmcl.2022.128949 doi /cbs_pica/cbs_olc/import_discovery/elsevier/einzuspielen/GBV00000000001878.pica (DE-627)ELV058702350 (ELSEVIER)S0960-894X(22)00425-5 DE-627 ger DE-627 rakwb eng 630 VZ 22 ssgn 46.00 bkl Wang, Ze verfasserin aut Ultrasensitive, rapid, and highly specific detection of microRNAs based on PER-CRISPR/CAS 2022transfer abstract nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier Abnormal microRNA (miRNA) expression levels are confirmed as diagnostic biomarkers of the emergence and development of diseases. In this study, we developed a fluorescence biosensor for detecting miRNAs based on double amplification reactions with the primer exchange reaction (PER) and CRISPR/Cas12a. In the absence of target miRNA-21, PER hairpins remained locked by the protector strands and the primers did not extend. In the presence of target miRNA-21, the miRNA-21 bound to the guard sequence and exposed primer binding sites. Also, the closed PER hairpin was unlocked to specifically extend primers into single-stranded DNA (ssDNA) of unequal lengths. These ssDNAs of unequal lengths could activate the cleavage of a reporter by Cas12a, leading to an increase in detectable fluorescence signals. A large number of short nucleic acid fragments were amplified by PER-CRISPR multiple cycle cleavage fluorescent probes. Based on PER-combined CRISPR/Cas12a established dual signal amplification method was characterized by a low limit of detection of 10fM. The fluorescent biosensor for miRNA detection had the advantages of low detection cost, simple operation, and mobility, providing a very promising platform for the point-of-care testing of miRNA-21. Abnormal microRNA (miRNA) expression levels are confirmed as diagnostic biomarkers of the emergence and development of diseases. In this study, we developed a fluorescence biosensor for detecting miRNAs based on double amplification reactions with the primer exchange reaction (PER) and CRISPR/Cas12a. In the absence of target miRNA-21, PER hairpins remained locked by the protector strands and the primers did not extend. In the presence of target miRNA-21, the miRNA-21 bound to the guard sequence and exposed primer binding sites. Also, the closed PER hairpin was unlocked to specifically extend primers into single-stranded DNA (ssDNA) of unequal lengths. These ssDNAs of unequal lengths could activate the cleavage of a reporter by Cas12a, leading to an increase in detectable fluorescence signals. A large number of short nucleic acid fragments were amplified by PER-CRISPR multiple cycle cleavage fluorescent probes. Based on PER-combined CRISPR/Cas12a established dual signal amplification method was characterized by a low limit of detection of 10fM. The fluorescent biosensor for miRNA detection had the advantages of low detection cost, simple operation, and mobility, providing a very promising platform for the point-of-care testing of miRNA-21. miRNA-21 Elsevier Fluorescence Elsevier PER Elsevier CRISPR/Cas12a Elsevier Wei, Hongguo oth Bu, Shengjun oth Li, Xue oth Zhou, Hongyu oth Zhang, Wenhui oth Wan, Jiayu oth Enthalten in Elsevier Science Torrecillas, S. ELSEVIER Feeding European sea bass ( 2018 a Tetrahedron publication for rapid dissemination of preliminary communications on all aspects of bioorganic chemistry, medicinal chemistry, bioinorganic chemistry and related disciplines Amsterdam [u.a.] (DE-627)ELV000272361 volume:74 year:2022 day:15 month:10 pages:0 https://doi.org/10.1016/j.bmcl.2022.128949 Volltext GBV_USEFLAG_U GBV_ELV SYSFLAG_U 46.00 Tiermedizin: Allgemeines VZ AR 74 2022 15 1015 0 |
spelling |
10.1016/j.bmcl.2022.128949 doi /cbs_pica/cbs_olc/import_discovery/elsevier/einzuspielen/GBV00000000001878.pica (DE-627)ELV058702350 (ELSEVIER)S0960-894X(22)00425-5 DE-627 ger DE-627 rakwb eng 630 VZ 22 ssgn 46.00 bkl Wang, Ze verfasserin aut Ultrasensitive, rapid, and highly specific detection of microRNAs based on PER-CRISPR/CAS 2022transfer abstract nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier Abnormal microRNA (miRNA) expression levels are confirmed as diagnostic biomarkers of the emergence and development of diseases. In this study, we developed a fluorescence biosensor for detecting miRNAs based on double amplification reactions with the primer exchange reaction (PER) and CRISPR/Cas12a. In the absence of target miRNA-21, PER hairpins remained locked by the protector strands and the primers did not extend. In the presence of target miRNA-21, the miRNA-21 bound to the guard sequence and exposed primer binding sites. Also, the closed PER hairpin was unlocked to specifically extend primers into single-stranded DNA (ssDNA) of unequal lengths. These ssDNAs of unequal lengths could activate the cleavage of a reporter by Cas12a, leading to an increase in detectable fluorescence signals. A large number of short nucleic acid fragments were amplified by PER-CRISPR multiple cycle cleavage fluorescent probes. Based on PER-combined CRISPR/Cas12a established dual signal amplification method was characterized by a low limit of detection of 10fM. The fluorescent biosensor for miRNA detection had the advantages of low detection cost, simple operation, and mobility, providing a very promising platform for the point-of-care testing of miRNA-21. Abnormal microRNA (miRNA) expression levels are confirmed as diagnostic biomarkers of the emergence and development of diseases. In this study, we developed a fluorescence biosensor for detecting miRNAs based on double amplification reactions with the primer exchange reaction (PER) and CRISPR/Cas12a. In the absence of target miRNA-21, PER hairpins remained locked by the protector strands and the primers did not extend. In the presence of target miRNA-21, the miRNA-21 bound to the guard sequence and exposed primer binding sites. Also, the closed PER hairpin was unlocked to specifically extend primers into single-stranded DNA (ssDNA) of unequal lengths. These ssDNAs of unequal lengths could activate the cleavage of a reporter by Cas12a, leading to an increase in detectable fluorescence signals. A large number of short nucleic acid fragments were amplified by PER-CRISPR multiple cycle cleavage fluorescent probes. Based on PER-combined CRISPR/Cas12a established dual signal amplification method was characterized by a low limit of detection of 10fM. The fluorescent biosensor for miRNA detection had the advantages of low detection cost, simple operation, and mobility, providing a very promising platform for the point-of-care testing of miRNA-21. miRNA-21 Elsevier Fluorescence Elsevier PER Elsevier CRISPR/Cas12a Elsevier Wei, Hongguo oth Bu, Shengjun oth Li, Xue oth Zhou, Hongyu oth Zhang, Wenhui oth Wan, Jiayu oth Enthalten in Elsevier Science Torrecillas, S. ELSEVIER Feeding European sea bass ( 2018 a Tetrahedron publication for rapid dissemination of preliminary communications on all aspects of bioorganic chemistry, medicinal chemistry, bioinorganic chemistry and related disciplines Amsterdam [u.a.] (DE-627)ELV000272361 volume:74 year:2022 day:15 month:10 pages:0 https://doi.org/10.1016/j.bmcl.2022.128949 Volltext GBV_USEFLAG_U GBV_ELV SYSFLAG_U 46.00 Tiermedizin: Allgemeines VZ AR 74 2022 15 1015 0 |
allfields_unstemmed |
10.1016/j.bmcl.2022.128949 doi /cbs_pica/cbs_olc/import_discovery/elsevier/einzuspielen/GBV00000000001878.pica (DE-627)ELV058702350 (ELSEVIER)S0960-894X(22)00425-5 DE-627 ger DE-627 rakwb eng 630 VZ 22 ssgn 46.00 bkl Wang, Ze verfasserin aut Ultrasensitive, rapid, and highly specific detection of microRNAs based on PER-CRISPR/CAS 2022transfer abstract nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier Abnormal microRNA (miRNA) expression levels are confirmed as diagnostic biomarkers of the emergence and development of diseases. In this study, we developed a fluorescence biosensor for detecting miRNAs based on double amplification reactions with the primer exchange reaction (PER) and CRISPR/Cas12a. In the absence of target miRNA-21, PER hairpins remained locked by the protector strands and the primers did not extend. In the presence of target miRNA-21, the miRNA-21 bound to the guard sequence and exposed primer binding sites. Also, the closed PER hairpin was unlocked to specifically extend primers into single-stranded DNA (ssDNA) of unequal lengths. These ssDNAs of unequal lengths could activate the cleavage of a reporter by Cas12a, leading to an increase in detectable fluorescence signals. A large number of short nucleic acid fragments were amplified by PER-CRISPR multiple cycle cleavage fluorescent probes. Based on PER-combined CRISPR/Cas12a established dual signal amplification method was characterized by a low limit of detection of 10fM. The fluorescent biosensor for miRNA detection had the advantages of low detection cost, simple operation, and mobility, providing a very promising platform for the point-of-care testing of miRNA-21. Abnormal microRNA (miRNA) expression levels are confirmed as diagnostic biomarkers of the emergence and development of diseases. In this study, we developed a fluorescence biosensor for detecting miRNAs based on double amplification reactions with the primer exchange reaction (PER) and CRISPR/Cas12a. In the absence of target miRNA-21, PER hairpins remained locked by the protector strands and the primers did not extend. In the presence of target miRNA-21, the miRNA-21 bound to the guard sequence and exposed primer binding sites. Also, the closed PER hairpin was unlocked to specifically extend primers into single-stranded DNA (ssDNA) of unequal lengths. These ssDNAs of unequal lengths could activate the cleavage of a reporter by Cas12a, leading to an increase in detectable fluorescence signals. A large number of short nucleic acid fragments were amplified by PER-CRISPR multiple cycle cleavage fluorescent probes. Based on PER-combined CRISPR/Cas12a established dual signal amplification method was characterized by a low limit of detection of 10fM. The fluorescent biosensor for miRNA detection had the advantages of low detection cost, simple operation, and mobility, providing a very promising platform for the point-of-care testing of miRNA-21. miRNA-21 Elsevier Fluorescence Elsevier PER Elsevier CRISPR/Cas12a Elsevier Wei, Hongguo oth Bu, Shengjun oth Li, Xue oth Zhou, Hongyu oth Zhang, Wenhui oth Wan, Jiayu oth Enthalten in Elsevier Science Torrecillas, S. ELSEVIER Feeding European sea bass ( 2018 a Tetrahedron publication for rapid dissemination of preliminary communications on all aspects of bioorganic chemistry, medicinal chemistry, bioinorganic chemistry and related disciplines Amsterdam [u.a.] (DE-627)ELV000272361 volume:74 year:2022 day:15 month:10 pages:0 https://doi.org/10.1016/j.bmcl.2022.128949 Volltext GBV_USEFLAG_U GBV_ELV SYSFLAG_U 46.00 Tiermedizin: Allgemeines VZ AR 74 2022 15 1015 0 |
allfieldsGer |
10.1016/j.bmcl.2022.128949 doi /cbs_pica/cbs_olc/import_discovery/elsevier/einzuspielen/GBV00000000001878.pica (DE-627)ELV058702350 (ELSEVIER)S0960-894X(22)00425-5 DE-627 ger DE-627 rakwb eng 630 VZ 22 ssgn 46.00 bkl Wang, Ze verfasserin aut Ultrasensitive, rapid, and highly specific detection of microRNAs based on PER-CRISPR/CAS 2022transfer abstract nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier Abnormal microRNA (miRNA) expression levels are confirmed as diagnostic biomarkers of the emergence and development of diseases. In this study, we developed a fluorescence biosensor for detecting miRNAs based on double amplification reactions with the primer exchange reaction (PER) and CRISPR/Cas12a. In the absence of target miRNA-21, PER hairpins remained locked by the protector strands and the primers did not extend. In the presence of target miRNA-21, the miRNA-21 bound to the guard sequence and exposed primer binding sites. Also, the closed PER hairpin was unlocked to specifically extend primers into single-stranded DNA (ssDNA) of unequal lengths. These ssDNAs of unequal lengths could activate the cleavage of a reporter by Cas12a, leading to an increase in detectable fluorescence signals. A large number of short nucleic acid fragments were amplified by PER-CRISPR multiple cycle cleavage fluorescent probes. Based on PER-combined CRISPR/Cas12a established dual signal amplification method was characterized by a low limit of detection of 10fM. The fluorescent biosensor for miRNA detection had the advantages of low detection cost, simple operation, and mobility, providing a very promising platform for the point-of-care testing of miRNA-21. Abnormal microRNA (miRNA) expression levels are confirmed as diagnostic biomarkers of the emergence and development of diseases. In this study, we developed a fluorescence biosensor for detecting miRNAs based on double amplification reactions with the primer exchange reaction (PER) and CRISPR/Cas12a. In the absence of target miRNA-21, PER hairpins remained locked by the protector strands and the primers did not extend. In the presence of target miRNA-21, the miRNA-21 bound to the guard sequence and exposed primer binding sites. Also, the closed PER hairpin was unlocked to specifically extend primers into single-stranded DNA (ssDNA) of unequal lengths. These ssDNAs of unequal lengths could activate the cleavage of a reporter by Cas12a, leading to an increase in detectable fluorescence signals. A large number of short nucleic acid fragments were amplified by PER-CRISPR multiple cycle cleavage fluorescent probes. Based on PER-combined CRISPR/Cas12a established dual signal amplification method was characterized by a low limit of detection of 10fM. The fluorescent biosensor for miRNA detection had the advantages of low detection cost, simple operation, and mobility, providing a very promising platform for the point-of-care testing of miRNA-21. miRNA-21 Elsevier Fluorescence Elsevier PER Elsevier CRISPR/Cas12a Elsevier Wei, Hongguo oth Bu, Shengjun oth Li, Xue oth Zhou, Hongyu oth Zhang, Wenhui oth Wan, Jiayu oth Enthalten in Elsevier Science Torrecillas, S. ELSEVIER Feeding European sea bass ( 2018 a Tetrahedron publication for rapid dissemination of preliminary communications on all aspects of bioorganic chemistry, medicinal chemistry, bioinorganic chemistry and related disciplines Amsterdam [u.a.] (DE-627)ELV000272361 volume:74 year:2022 day:15 month:10 pages:0 https://doi.org/10.1016/j.bmcl.2022.128949 Volltext GBV_USEFLAG_U GBV_ELV SYSFLAG_U 46.00 Tiermedizin: Allgemeines VZ AR 74 2022 15 1015 0 |
allfieldsSound |
10.1016/j.bmcl.2022.128949 doi /cbs_pica/cbs_olc/import_discovery/elsevier/einzuspielen/GBV00000000001878.pica (DE-627)ELV058702350 (ELSEVIER)S0960-894X(22)00425-5 DE-627 ger DE-627 rakwb eng 630 VZ 22 ssgn 46.00 bkl Wang, Ze verfasserin aut Ultrasensitive, rapid, and highly specific detection of microRNAs based on PER-CRISPR/CAS 2022transfer abstract nicht spezifiziert zzz rdacontent nicht spezifiziert z rdamedia nicht spezifiziert zu rdacarrier Abnormal microRNA (miRNA) expression levels are confirmed as diagnostic biomarkers of the emergence and development of diseases. In this study, we developed a fluorescence biosensor for detecting miRNAs based on double amplification reactions with the primer exchange reaction (PER) and CRISPR/Cas12a. In the absence of target miRNA-21, PER hairpins remained locked by the protector strands and the primers did not extend. In the presence of target miRNA-21, the miRNA-21 bound to the guard sequence and exposed primer binding sites. Also, the closed PER hairpin was unlocked to specifically extend primers into single-stranded DNA (ssDNA) of unequal lengths. These ssDNAs of unequal lengths could activate the cleavage of a reporter by Cas12a, leading to an increase in detectable fluorescence signals. A large number of short nucleic acid fragments were amplified by PER-CRISPR multiple cycle cleavage fluorescent probes. Based on PER-combined CRISPR/Cas12a established dual signal amplification method was characterized by a low limit of detection of 10fM. The fluorescent biosensor for miRNA detection had the advantages of low detection cost, simple operation, and mobility, providing a very promising platform for the point-of-care testing of miRNA-21. Abnormal microRNA (miRNA) expression levels are confirmed as diagnostic biomarkers of the emergence and development of diseases. In this study, we developed a fluorescence biosensor for detecting miRNAs based on double amplification reactions with the primer exchange reaction (PER) and CRISPR/Cas12a. In the absence of target miRNA-21, PER hairpins remained locked by the protector strands and the primers did not extend. In the presence of target miRNA-21, the miRNA-21 bound to the guard sequence and exposed primer binding sites. Also, the closed PER hairpin was unlocked to specifically extend primers into single-stranded DNA (ssDNA) of unequal lengths. These ssDNAs of unequal lengths could activate the cleavage of a reporter by Cas12a, leading to an increase in detectable fluorescence signals. A large number of short nucleic acid fragments were amplified by PER-CRISPR multiple cycle cleavage fluorescent probes. Based on PER-combined CRISPR/Cas12a established dual signal amplification method was characterized by a low limit of detection of 10fM. The fluorescent biosensor for miRNA detection had the advantages of low detection cost, simple operation, and mobility, providing a very promising platform for the point-of-care testing of miRNA-21. miRNA-21 Elsevier Fluorescence Elsevier PER Elsevier CRISPR/Cas12a Elsevier Wei, Hongguo oth Bu, Shengjun oth Li, Xue oth Zhou, Hongyu oth Zhang, Wenhui oth Wan, Jiayu oth Enthalten in Elsevier Science Torrecillas, S. ELSEVIER Feeding European sea bass ( 2018 a Tetrahedron publication for rapid dissemination of preliminary communications on all aspects of bioorganic chemistry, medicinal chemistry, bioinorganic chemistry and related disciplines Amsterdam [u.a.] (DE-627)ELV000272361 volume:74 year:2022 day:15 month:10 pages:0 https://doi.org/10.1016/j.bmcl.2022.128949 Volltext GBV_USEFLAG_U GBV_ELV SYSFLAG_U 46.00 Tiermedizin: Allgemeines VZ AR 74 2022 15 1015 0 |
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In this study, we developed a fluorescence biosensor for detecting miRNAs based on double amplification reactions with the primer exchange reaction (PER) and CRISPR/Cas12a. In the absence of target miRNA-21, PER hairpins remained locked by the protector strands and the primers did not extend. In the presence of target miRNA-21, the miRNA-21 bound to the guard sequence and exposed primer binding sites. Also, the closed PER hairpin was unlocked to specifically extend primers into single-stranded DNA (ssDNA) of unequal lengths. These ssDNAs of unequal lengths could activate the cleavage of a reporter by Cas12a, leading to an increase in detectable fluorescence signals. A large number of short nucleic acid fragments were amplified by PER-CRISPR multiple cycle cleavage fluorescent probes. Based on PER-combined CRISPR/Cas12a established dual signal amplification method was characterized by a low limit of detection of 10fM. 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Ultrasensitive, rapid, and highly specific detection of microRNAs based on PER-CRISPR/CAS |
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Abnormal microRNA (miRNA) expression levels are confirmed as diagnostic biomarkers of the emergence and development of diseases. In this study, we developed a fluorescence biosensor for detecting miRNAs based on double amplification reactions with the primer exchange reaction (PER) and CRISPR/Cas12a. In the absence of target miRNA-21, PER hairpins remained locked by the protector strands and the primers did not extend. In the presence of target miRNA-21, the miRNA-21 bound to the guard sequence and exposed primer binding sites. Also, the closed PER hairpin was unlocked to specifically extend primers into single-stranded DNA (ssDNA) of unequal lengths. These ssDNAs of unequal lengths could activate the cleavage of a reporter by Cas12a, leading to an increase in detectable fluorescence signals. A large number of short nucleic acid fragments were amplified by PER-CRISPR multiple cycle cleavage fluorescent probes. Based on PER-combined CRISPR/Cas12a established dual signal amplification method was characterized by a low limit of detection of 10fM. The fluorescent biosensor for miRNA detection had the advantages of low detection cost, simple operation, and mobility, providing a very promising platform for the point-of-care testing of miRNA-21. |
abstractGer |
Abnormal microRNA (miRNA) expression levels are confirmed as diagnostic biomarkers of the emergence and development of diseases. In this study, we developed a fluorescence biosensor for detecting miRNAs based on double amplification reactions with the primer exchange reaction (PER) and CRISPR/Cas12a. In the absence of target miRNA-21, PER hairpins remained locked by the protector strands and the primers did not extend. In the presence of target miRNA-21, the miRNA-21 bound to the guard sequence and exposed primer binding sites. Also, the closed PER hairpin was unlocked to specifically extend primers into single-stranded DNA (ssDNA) of unequal lengths. These ssDNAs of unequal lengths could activate the cleavage of a reporter by Cas12a, leading to an increase in detectable fluorescence signals. A large number of short nucleic acid fragments were amplified by PER-CRISPR multiple cycle cleavage fluorescent probes. Based on PER-combined CRISPR/Cas12a established dual signal amplification method was characterized by a low limit of detection of 10fM. The fluorescent biosensor for miRNA detection had the advantages of low detection cost, simple operation, and mobility, providing a very promising platform for the point-of-care testing of miRNA-21. |
abstract_unstemmed |
Abnormal microRNA (miRNA) expression levels are confirmed as diagnostic biomarkers of the emergence and development of diseases. In this study, we developed a fluorescence biosensor for detecting miRNAs based on double amplification reactions with the primer exchange reaction (PER) and CRISPR/Cas12a. In the absence of target miRNA-21, PER hairpins remained locked by the protector strands and the primers did not extend. In the presence of target miRNA-21, the miRNA-21 bound to the guard sequence and exposed primer binding sites. Also, the closed PER hairpin was unlocked to specifically extend primers into single-stranded DNA (ssDNA) of unequal lengths. These ssDNAs of unequal lengths could activate the cleavage of a reporter by Cas12a, leading to an increase in detectable fluorescence signals. A large number of short nucleic acid fragments were amplified by PER-CRISPR multiple cycle cleavage fluorescent probes. Based on PER-combined CRISPR/Cas12a established dual signal amplification method was characterized by a low limit of detection of 10fM. The fluorescent biosensor for miRNA detection had the advantages of low detection cost, simple operation, and mobility, providing a very promising platform for the point-of-care testing of miRNA-21. |
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Ultrasensitive, rapid, and highly specific detection of microRNAs based on PER-CRISPR/CAS |
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