Highly Sensitive CRISPR/Cas12a-Based Fluorescence Detection of Porcine Reproductive and Respiratory Syndrome Virus
| Autor: | Siyuan Liu, Yuying Liao, Yalan Yang, Dagang Tao, Peng Yang, Shouzhang Sun, Yonggang Liu, Yijie Tang, Zhonglin Tang, Shengsong Xie, Bin Chen, Yulan Zhao |
|---|---|
| Rok vydání: | 2021 |
| Předmět: |
viruses
animal diseases Biomedical Engineering Recombinase Polymerase Amplification DNA Single-Stranded Real-Time Polymerase Chain Reaction Biochemistry Genetics and Molecular Biology (miscellaneous) Virus Fluorescence Genes Reporter Limit of Detection CRISPR Porcine respiratory and reproductive syndrome virus Polymerase biology Reverse Transcriptase Polymerase Chain Reaction virus diseases General Medicine respiratory system Porcine reproductive and respiratory syndrome virus biology.organism_classification Virology Highly sensitive Reverse transcription polymerase chain reaction biology.protein RNA Viral CRISPR-Cas Systems |
| Zdroj: | ACS synthetic biology. 10(10) |
| ISSN: | 2161-5063 |
| Popis: | Porcine reproductive and respiratory syndrome (PRRS) is an economically important disease of swine that is caused by PRRS virus (PRRSV). In this study, we established a fluorescence assay for highly sensitive detection of PRRSV through integration of the reverse transcription-recombinase polymerase amplification (RT-RPA)-coupled Cas12a system with an optical property of single stranded DNA-fluorescently quenched (ssDNA-FQ) reporter. This technique can achieve isothermal and visual detection of PRRSV in 25 min. In particular, the assay reaction can be completed in a single tube. The limit of sensitivity for PRRSV detection was single copy without cross-reactivity of other porcine viruses. Correlation between 11 PRRSV clinical samples measured by the quantitative reverse transcription polymerase chain reaction (RT-qPCR) and CRISPR/Cas12a assay was determined; the result showed that our results were highly accurate. To sum up, this study developed a visual, sensitive, and specific method of nucleic acid detection based on a CRISPR-Cas12a technique for the on-site detection of PRRSV. |
| Databáze: | OpenAIRE |
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