Complete genome characterisation of a novel coronavirus associated with severe human respiratory disease in Wuhan, China

Fan Wu; Su Zhao; Bin Yu; Yan-Mei Chen; Wen Wang; Yi Hu; Zhi-Gang Song; Zhao-Wu Tao; Jun-Hua Tian; Yuan-Yuan Pei; Ming-Li Yuan; Yu-Ling Zhang; Fa-Hui Dai; Yi Liu; Qi-Min Wang; Jiao-Jiao Zheng; Lin Xu; Edward C. Holmes; Yong-Zhen Zhang

The copyright holder for this preprint (which was not peer-reviewed) is the . https://doi.org/10.1101/2020.01.24.919183 doi: bioRxiv preprint a meta-transcriptomic library was constructed for pair-end (150 bp) sequencing using an 91 Illumina MiniSeq as previously described [4] [5] [6] [7] . In total, we generated 56,565,928 sequence 92 reads that were de novo assembled and screened for potential aetiologic agents. Of the 93 384,096 contigs assembled by Megahit 8 , the longest (30,474 nucleotides [nt]) had high 94 abundance and was closely related to a bat SARS-like coronavirus isolate -bat-SL-CoVZC45 95 (GenBank Accession MG772933) -previously sampled in China, with a nt identity of 89.1% 96 (Table S1 and S2). The genome sequence of this novel virus, as well as its termini, were 97 determined and confirmed by RT-PCR 9 and 5'/3' RACE kits (TaKaRa), respectively. This 98 new virus was designated as WH-Human 1 coronavirus (WHCV) (and has also been referred 99 to as '2019-nCoV') and its whole genome sequence (29,903 nt) has been assigned GenBank 100 accession number MN908947. Remapping the RNA-seq data against the complete genome of 101 WHCV resulted in an assembly of 123,613 reads, providing 99.99% genome coverage at a 102 mean depth of 6.04X (range: 0.01X -78.84X) ( Figure S2 ). The viral load in the BALF sample 103 was estimated by quantitative PCR (qPCR) to be 3.95×10 8 copies/mL ( Figure S3 ). 104 The viral genome organization of WHCV was characterized by sequence alignment 105 against two representative members of the genus Betacoronavirus: a human-origin 106 coronavirus (SARS-CoV Tor2, AY274119) and a bat-origin coronavirus 107 MG772933) (Figure 2 ). The un-translational regions (UTR) and open reading frame (ORF) of 108 WHCV were mapped based on this sequence alignment and ORF prediction. The WHCV 109 viral genome was similar to these two coronaviruses (Figure 2 and Table S3) , with a gene to SARS-CoV in that it carries a predicted ORF8 gene (366 nt in length) located between the 119 M and N ORF genes. The functions of WHCV ORFs were predicted based on those of known 120 coronaviruses and given in Table S5 . In a manner similar to SARS CoV Tor2, a leader 121 transcription regulatory sequence (TRS) and nine putative body TRSs could be readily 122 identified upstream of the 5' end of ORF, with the putative conserved TRS core sequence 123 appeared in two forms -the ACGAAC or CUAAAC (Table S6) .