Gene Discovery by EST Sequencing inToxoplasma gondiiReveals Sequences Restricted to the Apicomplexa

Abstract

To accelerate gene discovery and facilitate genetic mapping in the protozoan parasite Toxoplasma gondii, we have generated >7000 new ESTs from the 5Ј ends of randomly selected tachyzoite cDNAs. Comparison of the ESTs with the existing gene databases identified possible functions for more than 500 new T. gondii genes by virtue of sequence motifs shared with conserved protein families, including factors involved in transcription, translation, protein secretion, signal transduction, cytoskeleton organization, and metabolism. Despite this success in identifying new genes, more than 50% of the ESTs correspond to genes of unknown function, reflecting the divergent evolutionary status of this parasite. A newly recognized class of genes was identified based on its similarity to sequences known only from other members of the same phylum, therefore identifying sequences that are apparently restricted to the Apicomplexa. Such genes may underlie pathways common to this group of medically important parasites, therefore identifying potential targets for intervention.

The microneme protein MIC3 of Toxoplasma gondii is a secretory adhesin that binds to both the surface of the host cells and the surface of the parasite

“…5). Seven ESTs that matched this cDNA sequence were found in the T. gondii EST database (http:// genome.wustl.edu/est/toxoesthmpg.html) (Ajioka et al, 1998).…”

Gene Expression Profile of Human Bone Marrow Stromal Cells: High-Throughput Expressed Sequence Tag Sequencing Analysis

“…Based on the feasibility of EST analysis for gene discovery and pattern configuration in other cell types or organisms [31], we undertook a larger-scale EST project to sequence randomly isolated cDNAs from a HBMSC cDNA library. The …”

“…Although incomplete and not error-free, ESTs remain an effective means for novel gene discovery and generating biologically informative probes for mapping genes to chromosomes as sequence-tagged sites (STSs), for identifying mutations leading to heritable diseases, and for full-length cDNA cloning [21,26]. The advantages of this approach are as follows: (1) it can be pursued as a relatively inexpensive and rapid way to access many of the expressed genes of a cell or tissue type [29,30]; and (2) with the advent of highthroughput sequencing technology and an increased interest in genome-wide studies, it became clear that ESTs could be generated in sufficient numbers to provide a rapid means of gene discovery [28,31], especially for those searching for human disease genes or constructing physical maps of the human genome [32].Based on the feasibility of EST analysis for gene discovery and pattern configuration in other cell types or organisms [31], we undertook a larger-scale EST project to sequence randomly isolated cDNAs from a HBMSC cDNA library. The FIG.…”

The origins of apicomplexan sequence innovation

“…2; Carlton et al 2002;Gardner et al 2002Gardner et al , 2005Abrahamsen et al 2004;Xu et al 2004;Pain et al 2005). Furthermore, large expressed sequence tag (EST) collections have been generated for an additional nine species (Ajioka et al 1998;Howe 2001;Li et al 2003a;Cui et al 2005), which can be used to construct so-called ''partial genomes '' (Peregrin-Alvarez and Parkinson 2007;Wasmuth et al 2008). Studies of these data sets have provided insights into adaptations that relate to their parasitic ecology, such as the identification of expanded families of proteases in Plasmodium (Wu et al 2003).…”