Improving RT-LAMP detection of SARS-CoV-2 RNA through primer set selection and combination

Abstract

Reverse transcription loop-mediated isothermal amplification (RT-LAMP) has emerged as a viable molecular diagnostic method to expand the breadth and reach of nucleic acid testing, particularly for SARS-CoV-2 detection and surveillance. While rapidly growing in prominence, RT-LAMP remains a relatively new method compared to the standard RT-qPCR, and contribution to our body of knowledge on designing LAMP primer sets and assays can have significant impact on its utility and adoption. Here we select and evaluate 18 LAMP primer sets for SARS-CoV-2 previously identified as sensitive ones under various conditions, comparing their speed and sensitivity with two LAMP formulations each with 2 reaction temperatures. We find that both LAMP formulations have some effects on the speed and detection sensitivity and identify several primer sets with similar high sensitivity for different SARS-CoV-2 gene targets. Significantly we observe a consistent sensitivity enhancement by combining primer sets for different targets, confirming and building on earlier work to create a simple, general approach to building better and more sensitive RT-LAMP assays.