RNA sequencing by direct tagmentation of RNA/DNA hybrids

Lin, Dachuan; Fu, Yusi; Sun, Yue; Li, Jie; Liu, Lu; Yao, Jiacheng; Wang, Guanbo; Wu, Yaling; Lao, Kaiqin; Lee, Raymond W.; Zheng, Genhua; Xu, Jun; Oh, Juntaek; Wang, Dong; Xie, Xiaohui; Huang, Yanyi; Wang, Jianbin

doi:10.1073/pnas.1919800117

Cited by 102 publications

(76 citation statements)

References 42 publications

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“…Although the clinical verification has not been carried out yet, this technology, once proved, might be conducive to rapid diagnosis of the disease [43]. A research group of Peking University claimed to have developed a new method for rapid construction of transcriptome sequencing library of SHERRY, which is helpful for rapid sequencing of SARS-CoV-2 [44]. found that chloroquine has an immune-modulating activity and could effectively inhibit in this virus in vitro [46] .…”

Section: Diagnosis Of Sars-cov-2mentioning

confidence: 99%

Review of the 2019 novel coronavirus (SARS-CoV-2) based on current evidence

Wang

et al. 2020

International Journal of Antimicrobial Agents

981

815

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Section: Diagnosis Of Sars-cov-2mentioning

confidence: 99%

Review of the 2019 novel coronavirus (SARS-CoV-2) based on current evidence

Wang

et al. 2020

International Journal of Antimicrobial Agents

981

815

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“…Current real-time RT-PCR techniques are not convenient, rapid, robust and also not 100% accurate (high rate of false negative). Furthermore, these methods (SHERRY), which is helpful for rapid sequencing of SARS-CoV-2 (Di et al, 2020). They showed that Tn5 transposase, which randomly binds and cuts double-stranded DNA, can directly fragment and prime the RNA/DNA heteroduplexes generated by reverse transcription.…”

Section: Diagnostic Tools For the Covid-19mentioning

confidence: 99%

Genomic Diversity and Evolution, Diagnosis, Prevention, and Therapeutics of the Pandemic COVID-19 Disease

Hoque¹,

Chaudhury²,

Akanda³

et al. 2020

Preprint

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A novel coronavirus COVID-19 was first emerged in Wuhan city of Hubei Province in China in December 2019. The COVID-19, since then spreads to 213 countries and territories, and has become a pandemic. Genomic analyses have indicated that the virus, popularly named as corona, originated through a natural process and is probably not a purposefully manipulated laboratory construct. However, currently available data are not sufficient to precisely conclude the origin of this fearsome virus. Genome-wide annotation of thousands of genomes revealed that more than 1,407 nucleotide mutations and 722 amino acids replacements occurred at different positions of the SARS-CoV-2. The spike (S) glycoprotein of SARS-CoV-2 possesses a functional polybasic (furin) cleavage site at the S1-S2 boundary through the insertion of 12 nucleotides. It leads to the predicted acquisition of 3-O-linked glycan around the cleavage site. Although real-time RT-PCR methods targeting specific gene(s) have widely been used to diagnose the COVID-19 patients, however, recently developed more convenient, rapid, and specific diagnostic tools targeting IgM/IgG or newly developed plug and play methods should be available for resource-poor developing countries. Some drugs, vaccines and therapies have shown great promise in early trials, however, these candidates of preventive or therapeutic agents have to pass a long path of trials before being released for the practical application against COVID-19. This review updates current knowledge on origin, genomic evolution, development of the diagnostic tools and the preventive or therapeutic remedies of the COVID-19, and discusses on scopes for further research and effective management and surveillance of COVID-19.

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“…Currently, the most comprehensive strategy is the combination of metatranscriptomics profiling with post-library SARS-CoV-2 target enrichment( 6 ). However, in most conventional RNA-seq methods, the double-strand DNA ligation (dsDL) portion of the protocol is usually the most demanding on hands-on time and user technique( 7 ). When superimposed on the target enrichment process, these labor intensive and lengthy protocols become impractical for routine use in the clinic, much less for the timely monitoring of viral genetics and evolution on large volumes of samples during an outbreak.…”

Section: Introductionmentioning

confidence: 99%

“…Recently, we reported a new RNA-seq library construction strategy that aims to address some of these challenges: SHERRY avoids the problematic dsDL step in library construction by taking advantage of the newly discovered Tn5 tagmentation activity on RNA/DNA hybrids, to directly tag RNA/cDNA fragments with sequencing adapters ( 7 ). As such, SHERRY has minimal sample transfers and greatly reduced hands-on time, making it simple, robust, and suitable for inputs ranging from single cells to 200 ng total RNA.…”

Section: Introductionmentioning

confidence: 99%

MINERVA: A facile strategy for SARS-CoV-2 whole genome deep sequencing of clinical samples

Chen¹,

Lin

et al. 2020

Preprint

Self Cite

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32The novel coronavirus disease 2019 pandemic poses a serious public health 33 risk. Analyzing the genome of severe acute respiratory syndrome coronavirus 2 (SARS-34 CoV-2) from clinical samples is crucial for the understanding of viral spread and viral 35 evolution, as well as for vaccine development. Existing sample preparation methods for 36 viral genome sequencing are demanding on user technique and time, and thus not ideal 37 for time-sensitive clinical samples; these methods are also not optimized for high 38 performance on viral genomes. We have developed MetagenomIc RNA EnRichment 39 VirAl sequencing (MINERVA), a facile, practical, and robust approach for metagenomic 40 and deep viral sequencing from clinical samples. This approach uses direct tagmentation 41 of RNA/DNA hybrids using Tn5 transposase to greatly simplify the sequencing library 42 construction process, while subsequent targeted enrichment can generate viral genomes 43 with high sensitivity, coverage, and depth. We demonstrate the utility of MINERVA on 44 pharyngeal, sputum and stool samples collected from COVID-19 patients, successfully 45 obtaining both whole metatranscriptomes and complete high-depth high-coverage SARS-46 CoV-2 genomes from these clinical samples, with high yield and robustness. MINERVA 47 is compatible with clinical nucleic extracts containing carrier RNA. With a shortened 48 hands-on time from sample to virus-enriched sequencing-ready library, this rapid, 49 versatile, and clinic-friendly approach will facilitate monitoring of viral genetic variations 50 during outbreaks, both current and future. 51 52 53 Introduction 54As of April 24, 2020, the ongoing COVID-19 viral pandemic has affected more than 2.6 55 million people in over 200 countries and territories around the world, and has claimed 56 more than 180 thousand lives. Closely monitoring the genetic diversity and distribution of 57 viral strains at the population level is essential for epidemiological tracking, and for 58 understanding viral evolution and transmission; additionally examining the viral 59 heterogeneity within a single individual is imperative for diagnosis and treatment. The 60 disease-causing pathogen, severe acute respiratory syndrome coronavirus 2 (SARS-61 CoV-2), was identified from early disease cases and its draft genome sequenced within 62 weeks, thanks to the rapid responses from researchers around the world(1, 2). The initial 63 SARS-CoV-2 draft genome was obtained independently from the same early COVID-19 64 patient samples using various conventional RNA-seq sequencing library construction 65 methods. Although these library construction methods successfully generated a draft 66 genome, several drawbacks hinder the use of these methods for routine viral genome 67 sequencing from the surge of clinical samples during an outbreak. 68 69 One direct library construction approach which was used to generate the SARS-CoV-2 70 draft genome(1, 2) essentially captures each sample's entire metatranscriptome, in which 71 SARS-CoV-2 is just one species ...

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RNA sequencing by direct tagmentation of RNA/DNA hybrids

Cited by 102 publications

References 42 publications

Review of the 2019 novel coronavirus (SARS-CoV-2) based on current evidence

Review of the 2019 novel coronavirus (SARS-CoV-2) based on current evidence

Genomic Diversity and Evolution, Diagnosis, Prevention, and Therapeutics of the Pandemic COVID-19 Disease

MINERVA: A facile strategy for SARS-CoV-2 whole genome deep sequencing of clinical samples

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