High sensitivity detection of coronavirus SARS-CoV-2 using multiplex PCR and a multiplex-PCR-based metagenomic method

Chenyu Li; David N. Debruyne; Julia Spencer; Vidushi Kapoor; Lily Y. Liu; Bo Zhou; Lucie Lee; Rounak Feigelman; Grayson Burdon; Jeffrey Liu; Alejandra Oliva; Adam Borcherding; Hongdong Tan; Alexander E. Urban; Guoying Liu; Zhitong Liu

Furthermore, we also propose a metagenomic method for the potential identification of unknown pathogens. Metagenomic technology usually requires about 20 to 100 million of sequencing reads in order to detect minuscule numbers of targets embedded in the massive amounts of human background. However, our method appears to selectively amplify target sequences. This bias permits the obtention of around 16% of target bases by sequencing at a depth of about 1 million total reads. It is thus especially suitable for the detection of novel pathogens and highly genetically unstable pathogens, such as influenza viruses. The exact mechanism of deselecting human sequences is currently under investigation. The random hexamer preference, chromosomal structure, sequence composition, target length, circularity, methylation status, telomeric and centromeric regions, as well as the edge effect, may influence such outcome. Ultimately, the observed preferential amplification towards more "random" sequences could provide us with an advantageous edge for the continuous improvement of this method.