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10.1016/j.ebiom.2020.103036 http://scihub22266oqcxt.onion/10.1016/j.ebiom.2020.103036 33045467!7544594!33045467     free     free     free Deprecated: Implicit conversion from float 231.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 231.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 231.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 231.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534 Deprecated: Implicit conversion from float 231.6 to int loses precision in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 534       EBioMedicine 2020 ; 61 (ä): 103036 Nephropedia Template TP {{tp|p=33045467|t=2020. A CRISPR-Cas12a-based specific enhancer for more sensitive detection of SARS-CoV-2 infection |pdf=|usr=}}{{33045467}} gab.com Text A CRISPR-Cas12a-based specific enhancer for more sensitive detection of SARS-CoV-2 infection JO: EBioMedicine http://www.kidney.de/pget.php?&tw=1&pmid=33045467 #FOAvip BACKGROUND: Real-time reverse transcription-PCR (rRT-PCR) has been the most effective and widely implemented diagnostic technology since the beginning of the COVID-19 pandemic. However, fuzzy rRT-PCR readouts with high Ct values are frequently encountered, resulting in uncertainty in diagnosis. METHODS: A Specific Enhancer for PCR-amplified Nucleic Acid (SENA) was developed based on the Cas12a trans-cleavage activity, which is specifically triggered by the rRT-PCR amplicons of the SARS-CoV-2 Orf1ab (O) and N fragments. SENA was first characterized to determine its sensitivity and specificity, using a systematic titration experiment with pure SARS-CoV-2 RNA standards, and was then verified in several hospitals, employing a couple of commercial rRT-PCR kits and testing various clinical specimens under different scenarios. FINDINGS: The ratio (10 min/5 min) of fluorescence change (FC) with mixed SENA reaction (mix-FCratio) was defined for quantitative analysis of target O and N genes, and the Limit of Detection (LoD) of mix-FCratio with 95% confidence interval was 1.2</=1.6</=2.1. Totally, 295 clinical specimens were analyzed, among which 21 uncertain rRT-PCR cases as well as 4 false negative and 2 false positive samples were characterized by SENA and further verified by next-generation sequencing (NGS). The cut-off values for mix-FCratio were determined as 1.145 for positive and 1.068 for negative. INTERPRETATION: SENA increases both the sensitivity and the specificity of rRT-PCR, solving the uncertainty problem in COVID-19 diagnosis and thus providing a simple and low-cost companion diagnosis for combating the pandemic. FUNDING: Detailed funding information is available at the end of the manuscript. Twit Text FOAVip A CRISPR-Cas12a-based specific enhancer for more sensitive detection of SARS-CoV-2 infection ????EBioMedicine http://www.kidney.de/pget.php?&tw=1&pmid=33045467 #FOAvip Twit Text # Free Paper on # # # # # LINK http://www.kidney.de/pget.php?&tw=1&pmid=33045467 #FOAvip English Wikipedia <ref>{{Cite pmid|33045467}}</ref> A CRISPR-Cas12a-based specific enhancer for more sensitive detection of SARS-CoV-2 infection #MMPMID33045467 Huang W; Yu L; Wen D; Wei D; Sun Y; Zhao H; Ye Y; Chen W; Zhu Y; Wang L; Wang L; Wu W; Zhao Q; Xu Y; Gu D; Nie G; Zhu D; Guo Z; Ma X; Niu L; Huang Y; Liu Y; Peng B; Zhang R; Zhang X; Li D; Liu Y; Yang G; Liu L; Zhou Y; Wang Y; Hou T; Gao Q; Li W; Chen S; Hu X; Han M; Zheng H; Weng J; Cai Z; Zhang X; Song F; Zhao G; Wang J EBioMedicine 2020[Nov]; 61 (ä): 103036 PMID33045467show ga BACKGROUND: Real-time reverse transcription-PCR (rRT-PCR) has been the most effective and widely implemented diagnostic technology since the beginning of the COVID-19 pandemic. However, fuzzy rRT-PCR readouts with high Ct values are frequently encountered, resulting in uncertainty in diagnosis. METHODS: A Specific Enhancer for PCR-amplified Nucleic Acid (SENA) was developed based on the Cas12a trans-cleavage activity, which is specifically triggered by the rRT-PCR amplicons of the SARS-CoV-2 Orf1ab (O) and N fragments. SENA was first characterized to determine its sensitivity and specificity, using a systematic titration experiment with pure SARS-CoV-2 RNA standards, and was then verified in several hospitals, employing a couple of commercial rRT-PCR kits and testing various clinical specimens under different scenarios. FINDINGS: The ratio (10 min/5 min) of fluorescence change (FC) with mixed SENA reaction (mix-FCratio) was defined for quantitative analysis of target O and N genes, and the Limit of Detection (LoD) of mix-FCratio with 95% confidence interval was 1.2 |Bacterial Proteins/*metabolism[MESH] |Betacoronavirus/*genetics/isolation & purification[MESH] |COVID-19[MESH] |CRISPR-Associated Proteins/*metabolism[MESH] |Clustered Regularly Interspaced Short Palindromic Repeats/*genetics[MESH] |Coronavirus Infections/diagnosis/pathology/virology[MESH] |Coronavirus Nucleocapsid Proteins[MESH] |Endodeoxyribonucleases/*metabolism[MESH] |Humans[MESH] |Limit of Detection[MESH] |Nasal Cavity/virology[MESH] |Nucleic Acid Amplification Techniques/methods/standards[MESH] |Nucleocapsid Proteins/genetics/metabolism[MESH] |Pandemics[MESH] |Phosphoproteins[MESH] |Pneumonia, Viral/diagnosis/pathology/virology[MESH] |Polyproteins[MESH] |RNA, Viral/genetics/*metabolism[MESH] |Real-Time Polymerase Chain Reaction/*methods/standards[MESH] |Reference Standards[MESH] |SARS-CoV-2[MESH]A CRISPR-Cas12a-based specific enhancer for more sensitive detection o(epmc).pdf DeepDyve Pubget Overpricing