Thursday, October 28, 2010

A novel multifunctional oligonucleotide microarray for Toxoplasma gondii

BMC Genomics. 2010 Oct 25;11(1):603. [Epub ahead of print]

A novel multifunctional oligonucleotide microarray for Toxoplasma gondii

Bahl A, Davis PH, Behnke M, Dzierszinski F, Jagalur M, Chen F, Shanmugam D, White M, Kulp D, Roos DS.


BACKGROUND: Microarrays are invaluable tools for genome interrogation, SNP detection, and expression analysis, among other applications. Such broad capabilities would be of value to many pathogen research communities, although the development and use of genome-scale microarrays is often a prohibitively expensive undertaking.

RESULTS: Taking advantage of available genome sequences and annotation for Toxoplasma gondii (the causative agent of opportunistic illness in immunocompromised individuals) and Plasmodium falciparum (a related parasite responsible for severe human malaria), we designed a single oligonucleotide microarray capable of supporting a wide range of applications at relatively low cost, including genome-wide expression profiling for Toxoplasma, and single-nucleotide polymorphism (SNP)-based genotyping of both T. gondii and P. falciparum. Expression profiling of the three clonotypic lineages dominating T. gondii populations in North America and Europe provides a first comprehensive view of the parasite transcriptome, revealing that ~49% of all annotated genes are expressed in parasite tachyzoites (the acutely lytic stage responsible for pathogenesis). The relatively small genomes of P. falciparum (23 Mb) and T. gondii (65 Mb), and the ability to isolate DNA from haploid stages, coupled with a careful analysis of signals from overlapping probes, permits a novel design enabling high confidence genotyping using only four probes per SNP. This strategy permits resolution of recombination points in the clonal progeny of sexual crosses, allowing detailed mapping of phenotypic traits. Recent sequencing of additional T. gondii isolates identifies >620K new SNPs, including >11K that intersect with expression profiling probes, yielding additional markers for genotyping studies, and further validating the utility of a combined expression profiling/genotyping array design. Additional applications facilitating gene and transcript discovery, statistical analysis of expression profiles, etc. are also discussed.

CONCLUSIONS: In addition to providing an initial global view of the T. gondii transcriptome across major lineages and permitting detailed resolution of recombination points in a historical sexual cross, the multifunctional nature of this array also allowed opportunities to exploit probes for purposes beyond their intended use, enhancing analyses. This array is in widespread use by the T. gondii research community, and several aspects of the design strategy are likely to be useful for other pathogens.

PMID: 20974003 [PubMed - as supplied by publisher]

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