This is a tailed amplicon method similar to https:\/\/www.protocols.io\/view\/sars-cov-2-tailed-amplicon-illumina-sequencing-bipikdke but based on the shorter 275nt amplicons in https:\/\/www.protocols.io\/view\/covid-19-artic-v3-illumina-library-construction-an-bh4zj8x6. One modification from the Gohl trailed protocol is the use of partial TruSeq tails instead of Nextera tails and using two vs four multiplexed pools. The advantage of this method is the speed and low cost due to several reasons. Only two separate cDNA ARTIC PCR reactions are required.After the indexing PCR step using TruSeq indexing primers, equal volumes are pooled and a single tube bead cleanup is performed. These libraries only require PE150 sequencing.The workflow can be completed in under 8 hour. If a small number of samples are prepared, a MiSeq Nano or MIcro run can be used, for a total time from RNA to FASTQs in roughly 24 hours. Note that this primer design has a 15bp primer overlap region around 20.5kb that results in 15 bases that cannot be sequenced with this method. We are investigating other primers in this region to recover these bases.We have optimized this method to process samples in 384-well format using a Beckman Biomek i7 and a Labcyte Echo to minimize tip usage (three tips per sample). The procedure can also be performed manually at lower scale or in 96-well format, but you may need to increase the volume of the reactions for more accurate pipeting. A cost effective manner to purchase this oligo set is with IDT oPools. At list price, you can order the two pools for a roughly $200 each. This is enough to prepare 500 samples at our volumes. You can also list each oligo two in a pool to get twice the amount of oligo at twice the cost.