RT-LAMP-CRISPR-Cas13a technology as a promising diagnostic tool for the SARS-CoV-2 virus

At the end of 2019, the new coronavirus, SARS-CoV-2, began a pandemic that persists to date and which has caused more than 6.2 million deaths. In the last couple of years, researchers have made great efforts to develop a diagnostic technique that maintains high levels of sensitivity and specificity, since an accurate and early diagnosis is required to minimize the prevalence of SARS-CoV-2 infection. In this context, CRISPR-Cas systems are proposed as promising tools for development in diagnostic techniques due to their high specificity, highlighting that Cas13 endonuclease discriminates single nucleotide changes and displays a collateral activity against single stranded RNA molecules. With the aim of improve the sensitivity of the diagnosis, this technology is usually combined with isothermal pre-amplification reactions (SHERLOCK, DETECTR). Basing on this, we have developed an RT-LAMP-CRISPR-Cas13a for SARS-CoV-2 virus detection in nasopharyngeal samples without using RNA extraction kit that exhibited 100 % specificity and 83 % sensitivity, as well as a positive predictive value of 100 % and a negative predictive value of 100%, 81%, 79.1% and 66.7 % in 30 Ct and total Ct values, respectively. Importance During Covid19 crisis has driven the development innovative molecular diagnose including the CRISPR-Cas technology. This work we have performed a protocol working with RNA-extraction kit free samples, places RT-LAMP-CRISPR-Cas13a technology at the top of rapid and specific diagnostic methods for COVID19 due to the high levels of specificity (100%), sensitivity (83%), PPV (100%) and NPV (81% in high loads viral) obtained in clinical samples.