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Molecular Biology

The advent of genomics and bioinformatics and the wider availability of microbial genome sequences has inevitably demanded different approaches to pathogen research. The combination of gene sequence analysis (genomics) with RNA expression analysis (transcriptomics), through peptide sequencing and biochemical/immunological characterisation, interfaced with protein expression analysis (proteomics) provides powerful tools in the understanding of pathogen biology and disease pathogenesis.

Of course, the interpretation of results requires the mining of relevant gene/protein databases and genome sequences are now available for most of the bacterial (e.g. Chlamydia abortus, Chlamydia pecorum, Pasteurella multocida, Cornynebacterium pseudotuberculosis, Mycobacterium paratuberculosis) and viral (e.g. Orf virus) pathogens that are the focus of research at Moredun.

Where genomes are not readily available for eukaryotic organisms (such as for nematode parasites Haemonchus contortus and Teladorsagia circumcincta and the sheep scab mite, Psoroptes ovis) Expressed Sequence Tag (EST) datasets are available. But how is all this information exploited in our bid to develop effective disease control strategies?

  • Gene expression can be quantified using real-time PCR approaches, DNA microarray technology or Serial Analysis of Gene Expression (SAGE); high-throughput sequencing now offers a further means for acquiring transcriptomic profiles. These techniques identify genes being expressed by an organism or tissue, their relative abundance and identify upregulated or down regulated genes. Gene expression can be characterised in response to particular stimuli such as infection in a tissue. In parallel, the pattern of gene expression in the infecting organism can be defined during the disease process. This leads to a very detailed insight into host/pathogen interactions and results in the identification of many novel genes and pathways associated with infection and therefore possible targets for disease control mechanisms.
  • Protein analysis techniques such as ‘SOSPA’ (Sawn-Off Shotgun Protein Analysis) and 2-D gel electrophoresis combined with MALDI or LC-ESI-MS/MS and immunoproteomics, are used routinely in-house and can be used to allow the identification of immunogenic protein components in subcellular fractions (such as protective membrane fractions), bacterial factors that promote infection (i.e. virulence factors), as well as to identify differences in protein expression between wild-type and attenuated strains, thus identifying target candidate antigens that can be used in the development of novel diagnostic tools and vaccines for detection and intervention.

Most recently, collaborations are underway with the Sanger Institute to conduct deep genome sequencing, using ‘Next Generation’ technologies, of some of the pathogens under study, specifically Mycobacterium and Chlamydia spp. The use of such Next Generation genome sequencing is enabling an unprecedented insight into the variability of field strains, and will provide important information in our understanding of how these pathogens differ in their ability to induce disease and how the host responds to infection.

Central to all of these techniques/technologies are the Moredun Proteomics Facility and Bioinformatics Unit, both of which impact on the Molecular Biology Theme and are now well established within Moredun. The capabilities provided by each of these underpin the science conducted at Moredun, cutting across each of the Divisions.

Virtually every research group is utilising the technologies or services provided by the two facilities in their work, whether it is 2-D gel electrophoresis, MALDI or Shotgun analysis, database creation, genome annotation and comparisons, etc. The Molecular Biology theme aims to bring all of these aspects together by facilitating interactions between research groups in each of the divisions through meetings and seminars to discuss applications of techniques and the use of novel approaches to solve or address specific problems or questions, to troubleshooting problems, as well as presenting results and solutions. Workshops on specific areas of science relating to the Molecular Biology are also held.

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Printed from http://www.moredun.org.uk/research/expertise-%40-moredun/molecular-biology on 27/05/17 09:19:58 AM

Moredun is committed to promoting animal health and welfare through research and education and is recognized worldwide for its contribution to research into infectious diseases of farmed livestock.