**7. Molecular diagnostics**

COVID-19 Test of SARS-CoV-2 is a real-time reverse transcription polymerase chain-reaction (PCR) in upper or lower respiratory samples for the qualitative identification of nucleic acid (such as nasal, nasopharyngeal or oropharyngeal swabs, sputum, lower respiratory tract aspirates, bronchoalveolar lavage, nasopharyngeal wash/aspirate or a nasal aspirate) that is individually obtained from those suspected of COVID-19 by their healthcare provider [75]. The latest COVID-19 outbreak can be detected using qPCR, but insufficient possession of reagents and equipment has hindered the identification of diseases. To assist in making COVID-19 more effective in our diagnostics, a new protocol was suggested for the application of the CRISPR-based SHERLOCK technique for detecting COVID-9. COVID-19 objectives were identified between 20 and 200 aM (10–100 copies per input microlitre) with the use of synthetic COVID- 19 virus RNA fragments. The test can be performed starting with patient-purified RNA as used in qRT-PCR trials and read in less than an hour with a dipstick, without the need for complex instrumentation [58, 59]. GolayMetaMiner, an in-house software, has identified four different regions over 50 nucleotides for the SARS-CoV-2 genome with 96 SARS-CoV-2 and 104 non-SARS-CoV-2 coronaviral genomes. Primers were made to target the longest and previously not targeted nsp2 region and tailored as a reverse transcription-polymerase chain reaction (RT-PCR) test without a probe. The new COVID- 19-nsp2 assay had a detection limit (LOD) of 1.8 TCID5 mL and did not intensify any human coronavirus pathogens and respiratory viruses. The process threshold reproducibility (Cp) values have been adequate and overall imprecision (%CV) values have dropped far below 5%. The latest assay evaluation using 59 clinical samples from 14 reported cases demonstrated a 100% compliance with COVID-19-RdRp/Hel reference assay, which has been previously established. A COVID-19-nsp2, fast sensitive RT-PCR test was developed for SARS-CoV-2 [76].

## **8. Future perspectives of nucleic acid-based vaccines**

Since COVID-19 is new to humanity and the essence of defensive immune responses is incompletely understood, it is unknown which vaccination techniques are going to be most effective. Therefore, designing diverse vaccine platforms and methods in tandem is crucial. Indeed, researchers worldwide have been racing to produce COVID-19 vaccines since the epidemic started, with at least 166 vaccine candidates now in preclinical and clinical production (Draft landscape of COVID-19 candidate vaccines, 2020). A new pandemic vaccine developing framework has been suggested to address the immediate need for a vaccine, compacting the development period from 10 to 15 years to 1 to 2 years [77]. Recombinant plasmid DNA has been investigated as a vaccine model, although lately, mRNA has appeared as a promising platform. Six mRNA-based COVID-19 vaccines and four DNA-based COVID-19 vaccines are currently in clinical trials, with 27 such vaccines (16 mRNAbased and 11 DNA-based) undergoing preclinical production [78]. (Draft landscape of COVID-19 candidate vaccines, 2020). For protein translation and post-translational modifications, antigen-encoding mRNA encapsulated with a carrier such as lipid nanoparticles can be effectively conveyed in vivo into the cytoplasm of host cells, which is a plus over vaccines of the recombinant protein subunit. The mRNA vaccines are non - pathogenic and are synthesized without microbial molecules by in vitro transcription [79]. While no mRNA vaccine has been approved for human use yet, recent reports of influenza, rabies and Zika virus infections in animals support its promise in the covid-19 vaccine development race [80]. Plasmid DNA vaccines share many features, such as safety, ease of development and scalability, with mRNA vaccines, but with the differences of having poor immunogenic and having to be administered in several doses coupled with the addition of an adjuvant. This review provides valuable information that can be redirected to the purpose of working on these nucleic acid-based vaccines which provides a new propitious platform of vaccine production.
