Isolating Viral and Host RNA Sequences from Archival Material and Production of cDNA Libraries for High-Throughput DNA Sequencing.
Identifieur interne : 000B42 ( Main/Exploration ); précédent : 000B41; suivant : 000B43Isolating Viral and Host RNA Sequences from Archival Material and Production of cDNA Libraries for High-Throughput DNA Sequencing.
Auteurs : Yongli Xiao [États-Unis] ; Zong-Mei Sheng [États-Unis] ; Jeffery K. Taubenberger [États-Unis]Source :
- Current protocols in microbiology [ 1934-8533 ] ; 2015.
Descripteurs français
- KwdFr :
- ARN viral (génétique), ARN viral (isolement et purification), Anatomopathologie moléculaire (), Banque de gènes, Humains, Pandémie de grippe de 1918-1919, Sous-type H1N1 du virus de la grippe A (génétique), Sous-type H1N1 du virus de la grippe A (isolement et purification), Séquençage nucléotidique à haut débit.
- MESH :
- génétique : ARN viral, Sous-type H1N1 du virus de la grippe A.
- isolement et purification : ARN viral, Sous-type H1N1 du virus de la grippe A.
- Anatomopathologie moléculaire, Banque de gènes, Humains, Pandémie de grippe de 1918-1919, Séquençage nucléotidique à haut débit.
English descriptors
- KwdEn :
- MESH :
- chemical , genetics : RNA, Viral.
- genetics : Influenza A Virus, H1N1 Subtype.
- isolation & purification : Influenza A Virus, H1N1 Subtype, RNA, Viral.
- methods : Pathology, Molecular.
- Gene Library, High-Throughput Nucleotide Sequencing, Humans, Influenza Pandemic, 1918-1919.
Abstract
The vast majority of surgical biopsy and post-mortem tissue samples are formalin-fixed and paraffin-embedded (FFPE), but this process leads to RNA degradation that limits gene expression analysis. As an example, the viral RNA genome of the 1918 pandemic influenza A virus was previously determined in a 9-year effort by overlapping RT-PCR from post-mortem samples. Using the protocols described here, the full genome of the 1918 virus was determined at high coverage in one high-throughput sequencing run of a cDNA library derived from total RNA of a 1918 FFPE sample after duplex-specific nuclease treatments. This basic methodological approach should assist in the analysis of FFPE tissue samples isolated over the past century from a variety of infectious diseases.
DOI: 10.1002/9780471729259.mc01e08s37
PubMed: 26344216
Affiliations:
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Le document en format XML
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<term>Influenza A Virus, H1N1 Subtype (isolation & purification)</term>
<term>Influenza Pandemic, 1918-1919</term>
<term>Pathology, Molecular (methods)</term>
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<term>Banque de gènes</term>
<term>Humains</term>
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<front><div type="abstract" xml:lang="en">The vast majority of surgical biopsy and post-mortem tissue samples are formalin-fixed and paraffin-embedded (FFPE), but this process leads to RNA degradation that limits gene expression analysis. As an example, the viral RNA genome of the 1918 pandemic influenza A virus was previously determined in a 9-year effort by overlapping RT-PCR from post-mortem samples. Using the protocols described here, the full genome of the 1918 virus was determined at high coverage in one high-throughput sequencing run of a cDNA library derived from total RNA of a 1918 FFPE sample after duplex-specific nuclease treatments. This basic methodological approach should assist in the analysis of FFPE tissue samples isolated over the past century from a variety of infectious diseases.</div>
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