The genome based detection of SARS-CoV-۲: Comparison of real-time-PCR and RT-LAMP

BACKGROUND AND ABJECTIVEOn December ۳۰, ۲۰۱۹, the coronavirus appeared and spread extremely rapidly, causingmillions of infections and deaths worldwide, and becoming a global pandemic. The mostpopular method for SARS-CoV-۲ detection is genome-based detection such as PCR basedthat they require expensive experimental equipments, controlled working environment andhigh-trained technician, which are often lacking in massive viral outbreaks. Therefore,another rapid and simple genome-based assay was evaluated for this pathogen. The aim ofthis work is to develop a new and sensitive loop test (RT-LAMP) using the E gene sequencefor rapid molecular diagnosis of patients, and compare it with the real time RT-PCR method.MATERIALS AND METHODSFor this purpose, ۱۸ samples of patients were collected by medical centers. The total RNA ofsamples was extracted according to the commercial kit (Roje). Following the initial setup, theRT-LAMP reaction was carried out using specific primers and the performance of thereaction was compared with the real-time PCR method. Also, the positive control construct(TA-E plasmid) was prepared by multiplying the relevant gene fragment and cloning into theTA vector.RESULTS AND DISCUSSIONThe sample examined by the real-time-PCR, the golden standard method, is evaluated with۱۰۰% conformity with the RT-LAMP method. That result shows the high sensitivity of theRT-LAMP method. It should be noted that in real-time-PCR test cycle threshold (CT) resultsless than ۳۰ are evaluated as positive. Also, the RT-LAMP assay has a sensitivity ofapproximately ۱۵ ng and ۱۱۲ pg for the E gene of SARS-CoV-۲ when using extracted totalRNA and TA-E plasmid, respectively.CONCLUSIONThe RT-LAMP method with equal sensitivity to real-time-PCR is an economical andisothermal method that can compensate for the disadvantages of the PCR method. Overall,this method can be used as a portable, rapid, and easy method for detecting SARS-CoV-۲ inthe field and clinical laboratories.

کلیدواژه ها:

SARS-CoV-۲ ، Genomic diagnosis ، RT-LAMP ، RT-PCR