Establishment Of Novel Coronavirus SARS-CoV-2 Nucleic Acid Diagnostic Method

The current situation of the new crown epidemic is still in the global epidemic.According to the current situation,the epidemic will continue for a relatively long time and will even coexist with humans in the future.Therefore,China may cause the new crown epidemic to spread in China at any time due to the introduction of the source of infection.Nucleic acid testing is currently the scientific basis for the diagnosis of new coronaviruses stipulated by governments around the world.Therefore,the establishment and optimization of high-efficiency extraction methods and accurate detection methods of new coronavirus nucleic acid have become the focus of industry experts and scholars.In this paper,three nucleic acid extraction methods and two nucleic acid diagnosis methods have been established,and clinical comparison and verification have been carried out.The experimental results show that among the three established methods for nucleic acid extraction and purification,the silica filter element has the highest purity for nucleic acid RNA extraction,followed by column centrifugation,and then magnetic bead method the established fluorescent PCR detection method can be detected when nucleic acid is 25copies/(?)L,one detection is equivalent to 250 copies,and the sensitivity is higher than the detection level of the market fluorescent quantitative PCR for a test of 300 copies or 500 copies.The established digital PCR detection method can detect the signal when the nucleic acid is 0.5 copies/(?)L,the sensitivity is significantly higher than that of fluorescent PCR,and the minimum detection limit is lower than that of fluorescent PCR.The clinical comparison of the fluorescent PCR detection method and the digital PCR detection method established in this paper shows that the established fluorescent PCR method and the commercial fluorescent PCR method of the third-party inspection agency are negative in the test of 384 samples,and the coincidence rate is 100%;The established digital PCR detection method and the commercial fluorescent PCR detection method of the First Hospital of Jilin University detected 7 positives in 197 environmental samples and 9positives in 136 human samples,with a coincidence rate of 100%,and using the established digital PCR detection method,5 cases of trace virus residue samples in the environment that cannot be detected by the fluorescence PCR detection method were detected,indicating that digital PCR is more sensitive than fluorescence PCR and can also detect trace virus residue.The nucleic acid extraction methods,fluorescent PCR detection methods,and new digital PCR detection methods established in this article provide early processing and sensitive and accurate detection for the development of suspected new coronavirus patients,early infection patients,and trace virus samples in the environment,and provide experiments and methodological basis.