tetano
Editor, Senior Moderator
Biomed Environ Sci. 2016 Jan;29(1):41-6. doi: 10.3967/bes2016.004.
[h=1]Evaluation of A Single-reaction Method for Whole Genome Sequencing of Influenza A Virus using Next Generation Sequencing.[/h] Zou XH[SUP]1[/SUP], Chen WB[SUP]1[/SUP], Zhao X[SUP]1[/SUP], Zhu WF[SUP]1[/SUP], Yang L[SUP]1[/SUP], Wang da Y[SUP]1[/SUP], Shu YL[SUP]1[/SUP].
[h=3]Author information[/h]
[h=3]Abstract[/h] [h=4]OBJECTIVE:[/h] To evaluate a single-reaction genome amplification method, the multisegment reverse transcription-PCR (M-RTPCR), for its sensitivity to full genome sequencing of influenza A virus, and the ability to differentiate mix-subtype virus, using the next generation sequencing (NGS) platform.
[h=4]METHODS:[/h] Virus genome copy was quantified and serially diluted to different titers, followed by amplification with the M-RTPCR method and sequencing on the NGS platform. Furthermore, we manually mixed two subtype viruses to different titer rate and amplified the mixed virus with the M-RTPCR protocol, followed by whole genome sequencing on the NGS platform. We also used clinical samples to test the method performance.
[h=4]RESULTS:[/h] The M-RTPCR method obtained complete genome of testing virus at 125 copies/reaction and determined the virus subtype at titer of 25 copies/reaction. Moreover, the two subtypes in the mixed virus could be discriminated, even though these two virus copies differed by 200-fold using this amplification protocol. The sensitivity of this protocol we detected using virus RNA was also confirmed with clinical samples containing low-titer virus.
[h=4]CONCLUSION:[/h] The M-RTPCR is a robust and sensitive amplification method for whole genome sequencing of influenza A virus using NGS platform.
Copyright ? 2016 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.
[h=4]KEYWORDS:[/h] Influenza A virus; NGS; Whole genome sequencing
PMID: 26822511 [PubMed - in process] Free full text
[h=1]Evaluation of A Single-reaction Method for Whole Genome Sequencing of Influenza A Virus using Next Generation Sequencing.[/h] Zou XH[SUP]1[/SUP], Chen WB[SUP]1[/SUP], Zhao X[SUP]1[/SUP], Zhu WF[SUP]1[/SUP], Yang L[SUP]1[/SUP], Wang da Y[SUP]1[/SUP], Shu YL[SUP]1[/SUP].
[h=3]Author information[/h]
[h=3]Abstract[/h] [h=4]OBJECTIVE:[/h] To evaluate a single-reaction genome amplification method, the multisegment reverse transcription-PCR (M-RTPCR), for its sensitivity to full genome sequencing of influenza A virus, and the ability to differentiate mix-subtype virus, using the next generation sequencing (NGS) platform.
[h=4]METHODS:[/h] Virus genome copy was quantified and serially diluted to different titers, followed by amplification with the M-RTPCR method and sequencing on the NGS platform. Furthermore, we manually mixed two subtype viruses to different titer rate and amplified the mixed virus with the M-RTPCR protocol, followed by whole genome sequencing on the NGS platform. We also used clinical samples to test the method performance.
[h=4]RESULTS:[/h] The M-RTPCR method obtained complete genome of testing virus at 125 copies/reaction and determined the virus subtype at titer of 25 copies/reaction. Moreover, the two subtypes in the mixed virus could be discriminated, even though these two virus copies differed by 200-fold using this amplification protocol. The sensitivity of this protocol we detected using virus RNA was also confirmed with clinical samples containing low-titer virus.
[h=4]CONCLUSION:[/h] The M-RTPCR is a robust and sensitive amplification method for whole genome sequencing of influenza A virus using NGS platform.
Copyright ? 2016 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.
[h=4]KEYWORDS:[/h] Influenza A virus; NGS; Whole genome sequencing
PMID: 26822511 [PubMed - in process] Free full text