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sars-cov‑2 and its omicron variants detection with rt-rpa -crispr/cas13a-based method at room temperature
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نویسنده
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li jia ,wang xiaojun ,chen liujie ,duan lili ,tan fenghua ,li kai ,hu zheng
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منبع
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reports of biochemistry and molecular biology - 2023 - دوره : 12 - شماره : 3 - صفحه:425 -437
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چکیده
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Background: the outbreak of severe acute respiratory syndrome coronavirus 2 (sars-cov-2) has triggered a global health crisis, with genetic mutations and evolution further creating uncertainty about epidemic risk. it is imperative to rapidly determine the nucleic acid sequence of sars-cov-2 and its variants to combat the coronavirus pandemic. our goal was to develop a rapid, room-temperature, point-of-care (poc) detection system to determine the nucleic acid sequences of sars-cov-2 isolates, especially omicron variants.methods: based on the conserved nucleotide sequence of sars-cov-2, bioinformatics software was used to analyze, design, and screen optimal enzymatic isothermal amplification primers and efficient crispr rnas (crrnas) of crispr/cas13a to the target sequences. reverse transcription-recombinase polymerase amplification (rt-rpa) was used to amplify the virus, and crispr/cas13a-crrna was used to cleave the sars-cov-2 target sequence. the sensitivity of nucleic acid detection was assessed by serial dilution of plasmid templates. all reactions were performed at room temperature.results: rt-rpa, combined with crispr/cas13a, can detect the sars-cov-2 with a minimum content of 102 copies/μl, and can effectively distinguish between the original strain and the omicron variant with a minimum limit of detection (lod) of 103 copies/μl.conclusion: the method developed in this study has potential application in clinical detection of sars-cov-2 and its omicron variants.
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کلیدواژه
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crispr/cas13a ,omicron variants ,rt-rpa ,sars-cov‑2
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آدرس
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xiangnan university, first people's hospital of chenzhou, first clinical college of xiangnan university, china. university of south china, translational medicine institute, first people's hospital of chenzhou, hengyang medical school, china, xiangnan university, first clinical college of xiangnan university, first people's hospital of chenzhou, china. first people's hospital of chenzhou, oncology department, china, xiangnan university, first clinical college of xiangnan university, first people's hospital of chenzhou, china. university of south china, translational medicine institute, first people's hospital of chenzhou, hengyang medical school, china, xiangnan university, first clinical college of xiangnan university, first people's hospital of chenzhou, china. university of south china, translational medicine institute, first people's hospital of chenzhou, hengyang medical school, china, xiangnan university, first clinical college of xiangnan university, first people's hospital of chenzhou, china. university of south china, translational medicine institute, first people's hospital of chenzhou, hengyang medical school, china, university of south china, translational medicine institute, first people's hospital of chenzhou, hengyang medical school, china. hunan university of chinese medicine, national engineering research center of personalized diagnostic and therapeutic technology, china, xiangnan university, first clinical college of xiangnan university, first people's hospital of chenzhou, china. university of south china, translational medicine institute, the first people's hospital of chenzhou, hengyang medical school, china
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پست الکترونیکی
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hu48005@163.com
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Authors
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