Tracking the evolution of the SARS coronavirus using high-throughput, high-density resequencing arrays.
Mutations in the SARS-Coronavirus (SARS-CoV) can alter its clinical presentation, and the study of its mutation patterns in human populations can facilitate contact tracing. Here, we describe the development and validation of an oligonucleotide resequencing array for interrogating the entire 30-kb SARS-CoV genome in a rapid, cost-effective fashion. Using this platform, we sequenced SARS-CoV genomes from Vero cell culture isolates of 12 patients and directly from four patient tissues. The sequence obtained from the array is highly reproducible, accurate (>99.99% accuracy) and capable of identifying known and novel variants of SARS-CoV. Notably, we applied this technology to a field specimen of probable SARS and rapidly deduced its infectious source. We demonstrate that array-based resequencing-by-hybridization is a fast, reliable, and economical alternative to capillary sequencing for obtaining SARS-CoV genomic sequence on a population scale, making this an ideal platform for the global monitoring of SARS-CoV and other small-genome pathogens.
Wong
Christopher W
CW
Genome Institute of Singapore, Singapore 138672, Republic of Singapore. wongc@gis.a-star.edu.sg
Albert
Thomas J
TJ
Vega
Vinsensius B
VB
Norton
Jason E
JE
Cutler
David J
DJ
Richmond
Todd A
TA
Stanton
Lawrence W
LW
Liu
Edison T
ET
Miller
Lance D
LD
D000818:Animals; D001482:Base Composition / Q000235:genetics; D020137:Base Pair Mismatch / Q000235:genetics; D002460:Cell Line; D002522:Chlorocebus aethiops / Q000235:genetics; D016384:Consensus Sequence / Q000235:genetics; D017934:Coronavirus / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification; D017931:DNA Primers / Q000235:genetics / Q000378:metabolism; D004279:DNA, Viral / Q000235:genetics / Q000378:metabolism; D019143:Evolution, Molecular; D016679:Genome, Viral; D006801:Humans; D009693:Nucleic Acid Hybridization / Q000235:genetics; D020411:Oligonucleotide Array Sequence Analysis; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism; D045473:Severe acute respiratory syndrome-related coronavirus / Q000145:classification / Q000235:genetics*; D017423:Sequence Analysis, RNA / Q000379:methods; D045169:Severe Acute Respiratory Syndrome / Q000175:diagnosis / Q000821:virology; D014709:Vero Cells / Q000821:virology
D017931:DNA Primers; D004279:DNA, Viral; D012367:RNA, Viral