Giulia Kennedy

Publications (2)2.86 Total impact

• Article: Use of consensus sequences for the design of high density resequencing microarrays: the influenza virus paradigm

  India Leclercq, Nicolas Berthet, Christophe Batéjat, Claudine Rousseaux, Philip Dickinson, Iain Old, Katherine Kong, Giulia Kennedy, Stewart Cole, Jean-Claude Manuguerra
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  ABSTRACT: Abstract Background A resequencing microarray called PathogenID v2.0 has been developed and used to explore various strategies of sequence selection for its design. The part dedicated to influenza viruses was based on consensus sequences specific for one gene generated from global alignments of a large number of influenza virus sequences available in databanks. Results For each HA (H1, H2, H3, H5, H7 and H9) and NA (N1, N2 and N7) molecular type chosen to be tested, 1 to 3 consensus sequences were computed and tiled on the microarray. A total of 12 influenza virus samples from different host origins (humans, pigs, horses and birds) and isolated over a period of about 50 years were used in this study. Influenza viruses were correctly identified, and in most cases with the accurate information of the time of their emergence. Conclusions PathogenID v2.0 microarray demonstrated its ability to type and subtype influenza viruses, often to the level of viral variants, with a minimum number of tiled sequences. This validated the strategy of using consensus sequences, which do not exist in nature, for our microarray design. The versatility, rapidity and high discriminatory power of the PathogenID v2.0 microarray could prove critical to detect and identify viral genome reassortment events resulting in a novel virus with epidemic or pandemic potential and therefore assist health authorities to make efficient decisions about patient treatment and outbreak management.
  BMC Genomics. 01/2010;
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  Available from: India Leclercq

  Article: Phi29 polymerase based random amplification of viral RNA as an alternative to random RT-PCR.

  Nicolas Berthet, Anita K Reinhardt, India Leclercq, Sven van Ooyen, Christophe Batéjat, Philip Dickinson, Rayna Stamboliyska, Iain G Old, Katherine A Kong, Laurent Dacheux, Hervé Bourhy, Giulia C Kennedy, Christian Korfhage, Stewart T Cole, Jean-Claude Manuguerra
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  ABSTRACT: Phi29 polymerase based amplification methods provides amplified DNA with minimal changes in sequence and relative abundance for many biomedical applications. RNA virus detection using microarrays, however, can present a challenge because phi29 DNA polymerase cannot amplify RNA nor small cDNA fragments (<2000 bases) obtained by reverse transcription of certain viral RNA genomes. Therefore, ligation of cDNA fragments is necessary prior phi29 polymerase based amplification. We adapted the QuantiTect Whole Transcriptome Kit (Qiagen) to our purposes and designated the method as Whole Transcriptome Amplification (WTA). WTA successfully amplified cDNA from a panel of RNA viruses representing the diversity of ribovirus genome sizes. We amplified a range of genome copy numbers from 15 to 4 x 10(7) using WTA, which yielded quantities of amplified DNA as high as 1.2 microg/microl or 10(10) target copies. The amplification factor varied between 10(9) and 10(6). We also demonstrated that co-amplification occurred when viral RNA was mixed with bacterial DNA. This is the first report in the scientific literature showing that a modified WGA (WTA) approach can be successfully applied to viral genomic RNA of all sizes. Amplifying viral RNA by WTA provides considerably better sensitivity and accuracy of detection compared to random RT-PCR.
  BMC Molecular Biology 10/2008; 9:77. · 2.86 Impact Factor