Publications des scientifiques de l'IRD

Guinoiseau T., Moreau A., Hohnadel G., Ngo-Giang-Huong Nicole, Brulard C., Vourc'h P., Goudeau A., Gaudy-Graffin C. (2017). Deep sequencing is an appropriate tool for the selection of unique Hepatitis C virus (HCV) variants after single genomic amplification. Plos One, 12 (3), p. e0174852 [11 p.]. ISSN 1932-6203.

Titre du document
Deep sequencing is an appropriate tool for the selection of unique Hepatitis C virus (HCV) variants after single genomic amplification
Année de publication
2017
Type de document
Article référencé dans le Web of Science WOS:000399175000044
Auteurs
Guinoiseau T., Moreau A., Hohnadel G., Ngo-Giang-Huong Nicole, Brulard C., Vourc'h P., Goudeau A., Gaudy-Graffin C.
Source
Plos One, 2017, 12 (3), p. e0174852 [11 p.] ISSN 1932-6203
Hepatitis C virus (HCV) evolves rapidly in a single host and circulates as a quasispecies wich is a complex mixture of genetically distinct virus's but closely related namely variants. To identify intra-individual diversity and investigate their functional properties in vitro, it is necessary to define their quasispecies composition and isolate the HCV variants. This is possible using single genome amplification (SGA). This technique, based on serially diluted cDNA to amplify a single cDNA molecule (clonal amplicon), has already been used to determine individual HCV diversity. In these studies, positive PCR reactions from SGA were directly sequenced using Sanger technology. The detection of non-clonal amplicons is necessary for excluding them to facilitate further functional analysis. Here, we compared Next Generation Sequencing (NGS) with De Novo assembly and Sanger sequencing for their ability to distinguish clonal and non-clonal amplicons after SGA on one plasma specimen. All amplicons (n = 42) classified as clonal by NGS were also classified as clonal by Sanger sequencing. No double peaks were seen on electropherograms for non-clonal amplicons with position-specific nucleotide variation below 15% by NGS. Altogether, NGS circumvented many of the difficulties encountered when using Sanger sequencing after SGA and is an appropriate tool to reliability select clonal amplicons for further functional studies.
Plan de classement
Entomologie médicale / Parasitologie / Virologie [052]
Description Géographique
THAILANDE
Localisation
Fonds IRD [F B010069512]
Identifiant IRD
fdi:010069512
Contact