1.Multiplex sequencing of SARS-Cov-2 genome directly from clinical samples using the Ion Personal Genome Machine (PGM)
Tan, K.K. ; Tiong, V. ; Tan, J.Y. ; Wong, J.E. ; Teoh, B.T. ; Abd-Jamil, J. ; Johari, J. ; Nor&rsquo ; e, S.S. ; Khor, C.S. ; Yaacob, C.N. ; Zulkifli, M.M.S. ; CheMatSeri, A. ; Mahfodz, N.H. ; Azizan, N.S. ; AbuBakar, S.
Tropical Biomedicine 2021;38(No.3):283-288
Various methods have been developed for rapid and high throughput full genome sequencing of SARS-CoV-2. Here, we described a protocol for targeted multiplex full genome sequencing of SARS-CoV-2 genomic RNA directly extracted from human nasopharyngeal swabs using the Ion Personal Genome Machine (PGM). This protocol involves concomitant amplification of 237 gene fragments encompassing the SARS-CoV-2 genome to increase the abundance and yield of viral specific sequencing reads. Five complete and one near-complete genome sequences of SARS-CoV-2 were generated with a single Ion PGM sequencing run. The sequence coverage analysis revealed two amplicons (positions 13 751-13 965 and 23 941-24 106), which consistently gave low sequencing read coverage in all isolates except 4Apr20-64Hu. We analyzed the potential primer binding sites within these low covered regions and noted that the 4Apr20-64-Hu possess C at positions 13 730 and 23 929, whereas the other isolates possess T at these positions. The genome nucleotide variations observed suggest that the naturally occurring variations present in the actively circulating SARS-CoV-2 strains affected the performance of the target enrichment panel of the Ion AmpliSeq™ SARS CoV 2 Research Panel. The possible impact of other genome nucleotide variations warrants further investigation, and an improved version of the Ion AmpliSeq™ SARS CoV 2 Research Panel, hence, should be considered.
Result Analysis
Print
Save
E-mail