Development and validation of an oligonucleotide microarray for immuno-inflammatory genes of ruminants

  • Craig Watkins
  • Annie McKellar
  • Kirsty Jensen
  • Abraham George
  • Doug Jones
  • Michael J. Sharp
  • Karen Stevenson
  • John Hopkins
Original Article

Abstract

This report describes the development of small DNA microarrays of fully defined genes suitable for projects requiring detailed analysis of gene expression in sheep and/or cattle. Two arrays have been developed; the first is a small reference microarray (RIGRA) that has been used to validate experimental design and methodology; the second, a larger array (RIGUA) containing probes for 516 ruminant immuno-inflammatory genes, each represented by non-overlapping 75mer oligonucleotides. Experiments used to validate this microarray were: (1) a comparison of gene expression profiles from sheep broncho-alveolar macrophages before and after in vitro activation with lipopolysaccharide (LPS), using the RIGRA; (2) the differential gene expression between five in vitro unstimulated sheep keratinocyte cultures; (3) LPS/interferon γ stimulated and unstimulated blood monocytes purified from Holstein-Friesians (Bos taurus) and Sahiwals (Bos indicus) cattle using the RIGUA. Real-time, quantitative RT-PCR was used to validate the gene expression profiles obtained with the RIGUA microarrays. The potential for using such an immunological tool in understanding the relative gene expression corresponding to immune-inflammatory responses of sheep and cattle is discussed.

Keywords

Ruminant Microarray Immuno-inflammatory Gene expression 

Abbreviations

aRNA

amplified RNA

OAM

ovine alveolar macrophages

RIGRA

Ruminant Immuno-inflammatory Gene Reference Array

RIGUA

Ruminant Immuno-inflammatory Gene Universal Array

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Copyright information

© Springer Science+Business Media B.V. 2008

Authors and Affiliations

  • Craig Watkins
    • 1
    • 3
  • Annie McKellar
    • 1
  • Kirsty Jensen
    • 2
  • Abraham George
    • 1
  • Doug Jones
    • 1
  • Michael J. Sharp
    • 1
    • 4
  • Karen Stevenson
    • 1
  • John Hopkins
    • 3
  1. 1.Moredun Research InstituteMidlothianUK
  2. 2.Roslin Institute & Royal (Dick) School of Veterinary StudiesUniversity of EdinburghMidlothianUK
  3. 3.Centre for Infectious Disease, Royal (Dick) School of Veterinary Studies (R(D)SVS)University of EdinburghEdinburghUK
  4. 4.Veterinary Laboratory AgencyMidlothianUK

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