The global outbreak of coronavirus disease 19(COVID-19),caused by severe acute respiratory syndrome coronavirus 2(SARS-Co V-2),has raised significant global apprehension.Developing a rapid,efficient,sensitive,and accu...The global outbreak of coronavirus disease 19(COVID-19),caused by severe acute respiratory syndrome coronavirus 2(SARS-Co V-2),has raised significant global apprehension.Developing a rapid,efficient,sensitive,and accurate point-of-care detection method is imperative for curbing SARS-Co V-2 transmission.Here,we screened a sequence,designed a set of highly sensitive loopmediated isothermal amplification primers(LAMP)and g RNA,and developed a user-friendly detection platform combining CRISPRCas12a and RT-LAMP technology to specifically detect SARS-Co V-2 and its 5 variants.Bioinformatics analysis and Cas12a-g RNA identification ensured sequence specificity,allowing us to identify SARS-Co V-2 mutations.We developed a method for the detection of SARSCoV-2 using these primers in combination with LAMP amplification and CRISPR-Cas12a technology.This method is designed to detect SARS-CoV-2(NC_045512),Alpha(B.1.1.7),Beta(B.1.351),Gamma(P.1),Delta(B.1.617.2)and Omicron(B.1.1.529).Additionally,it can differentiate SARS-CoV-2 from other coronaviruses.Quantitative analysis can be conducted by measuring fluorescence values,while qualitative analysis can be performed by observing fluorescence color point-of-care diagnosis changes with the naked eye.These results suggest that a set of novel sensitive LAMP primers and g RNA have been obtained to detect the extensive variants,and the RT-LAMPCRISPR-Cas12a platform significantly facilitates point-of-care diagnosis,thereby halting the spread of SARS-Co V-2,thus contributing to COVID-19 prevention and control.展开更多
Coronavirus disease(COVID-19)is a serious respiratory disease that spreads through the coronavirus globally.It soon became a pandemic after its appearance in 2019 and demanded new techniques for its identification and...Coronavirus disease(COVID-19)is a serious respiratory disease that spreads through the coronavirus globally.It soon became a pandemic after its appearance in 2019 and demanded new techniques for its identification and detection.Owing to this situation,RT-LAMP appears to be a novel method for the identification of COVID-19 because of its vast applications,including cost-effectiveness and time-saving.This research highlights the use of RT-LAMP,a more sensitive test than RT-PCR,for the assessment of SARS-CoV-2,the severe acute respiratory illness.To identify the spike(S)and NSP1 protein using RT-LAMP,170 total samples of coronavirus-suspected patients were served in this research.Health certifications and bioethical considerations were taken into consideration.After the sample was extracted from the patient's swabs,RNA was isolated,extracted,and purified.The response was then run on the RT-LAMP at the ideal temperature,and the outcomes could be observed with the unaided eye as they changed from pink to yellow.It is a simple method of determining if the test is positive or negative.For this purpose,both RT-LAMP and RT-PCR tests are used during these procedures.Genes linked with COVID-19 testing including S,nspl,and ORF are suited to coronavirus testing;they have 100%specificity and low sensitivity,but S has more specificity and sensitivity than nspl and ORF,respectively.Out of the 95 positive samples,89(93.68%)samples yielded favorable outcomes utilizing RT-LAMP,while 55 negative samples yielded 100%positive results.The present research demonstrates that RT-LAMP is less sensitive yet more selective for coronavirus detection.展开更多
基金Supported by the National Natural Sciences Foundation of China(52073022)the Fundamental Research Funds for the Central Universities of China and the Translational Medical Research Fund of Wuhan University Taikang Medical School(School of Basic Medical Sciences)the Key Laboratory of Environmental Pollution Monitoring and Disease Control(Guizhou Medical University)Ministry of Education(GMU-2022-HJZ)。
文摘The global outbreak of coronavirus disease 19(COVID-19),caused by severe acute respiratory syndrome coronavirus 2(SARS-Co V-2),has raised significant global apprehension.Developing a rapid,efficient,sensitive,and accurate point-of-care detection method is imperative for curbing SARS-Co V-2 transmission.Here,we screened a sequence,designed a set of highly sensitive loopmediated isothermal amplification primers(LAMP)and g RNA,and developed a user-friendly detection platform combining CRISPRCas12a and RT-LAMP technology to specifically detect SARS-Co V-2 and its 5 variants.Bioinformatics analysis and Cas12a-g RNA identification ensured sequence specificity,allowing us to identify SARS-Co V-2 mutations.We developed a method for the detection of SARSCoV-2 using these primers in combination with LAMP amplification and CRISPR-Cas12a technology.This method is designed to detect SARS-CoV-2(NC_045512),Alpha(B.1.1.7),Beta(B.1.351),Gamma(P.1),Delta(B.1.617.2)and Omicron(B.1.1.529).Additionally,it can differentiate SARS-CoV-2 from other coronaviruses.Quantitative analysis can be conducted by measuring fluorescence values,while qualitative analysis can be performed by observing fluorescence color point-of-care diagnosis changes with the naked eye.These results suggest that a set of novel sensitive LAMP primers and g RNA have been obtained to detect the extensive variants,and the RT-LAMPCRISPR-Cas12a platform significantly facilitates point-of-care diagnosis,thereby halting the spread of SARS-Co V-2,thus contributing to COVID-19 prevention and control.
文摘Coronavirus disease(COVID-19)is a serious respiratory disease that spreads through the coronavirus globally.It soon became a pandemic after its appearance in 2019 and demanded new techniques for its identification and detection.Owing to this situation,RT-LAMP appears to be a novel method for the identification of COVID-19 because of its vast applications,including cost-effectiveness and time-saving.This research highlights the use of RT-LAMP,a more sensitive test than RT-PCR,for the assessment of SARS-CoV-2,the severe acute respiratory illness.To identify the spike(S)and NSP1 protein using RT-LAMP,170 total samples of coronavirus-suspected patients were served in this research.Health certifications and bioethical considerations were taken into consideration.After the sample was extracted from the patient's swabs,RNA was isolated,extracted,and purified.The response was then run on the RT-LAMP at the ideal temperature,and the outcomes could be observed with the unaided eye as they changed from pink to yellow.It is a simple method of determining if the test is positive or negative.For this purpose,both RT-LAMP and RT-PCR tests are used during these procedures.Genes linked with COVID-19 testing including S,nspl,and ORF are suited to coronavirus testing;they have 100%specificity and low sensitivity,but S has more specificity and sensitivity than nspl and ORF,respectively.Out of the 95 positive samples,89(93.68%)samples yielded favorable outcomes utilizing RT-LAMP,while 55 negative samples yielded 100%positive results.The present research demonstrates that RT-LAMP is less sensitive yet more selective for coronavirus detection.
