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Nucleic Acids Research 2004 32(11):e90; doi:10.1093/nar/gnh088
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Published online 24 June 2004

Nucleic Acids Research, Vol. 32 No. 11 © Oxford University Press 2004; all rights reserved

Structural analysis of hepatitis C RNA genome using DNA microarrays

María Martell*, Carlos Briones1, Aránzazu de Vicente1, María Piron, Juan I. Esteban, Rafael Esteban, Jaime Guardia and Jordi Gómez

Laboratorio Medicina Interna-Hepatología, Hospital Vall d'Hebron, Barcelona, Spain and 1 Laboratorio de Evolución Molecular, Centro de Astrobiología, (CSIC-INTA), Madrid, Spain

* To whom correspondence should be addressed. Tel: +34 934894034; Fax: +34 934894032; Email: mmartell{at}vhebron.net

Received March 20, 2004; Revised May 20, 2004; Accepted June 3, 2004

Many studies have tried to identify specific nucleotide sequences in the quasispecies of hepatitis C virus (HCV) that determine resistance or sensitivity to interferon (IFN) therapy, unfortunately without conclusive results. Although viral proteins represent the most evident phenotype of the virus, genomic RNA sequences determine secondary and tertiary structures which are also part of the viral phenotype and can be involved in important biological roles. In this work, a method of RNA structure analysis has been developed based on the hybridization of labelled HCV transcripts to microarrays of complementary DNA oligonucleotides. Hybridizations were carried out at non-denaturing conditions, using appropriate temperature and buffer composition to allow binding to the immobilized probes of the RNA transcript without disturbing its secondary/tertiary structural motifs. Oligonucleotides printed onto the microarray covered the entire 5' non-coding region (5'NCR), the first three-quarters of the core region, the E2–NS2 junction and the first 400 nt of the NS3 region. We document the use of this methodology to analyse the structural degree of a large region of HCV genomic RNA in two genotypes associated with different responses to IFN treatment. The results reported here show different structural degree along the genome regions analysed, and differential hybridization patterns for distinct genotypes in NS2 and NS3 HCV regions.


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