Analyses of Expressed Sequence Tags from Chironomus riparius Using Pyrosequencing : Molecular Ecotoxicology Perspective

Abstract

OBJECTIVES: Chironomus riparius, a non-biting midge (Chironomidae, Diptera), is extensively used as a model organism in aquatic ecotoxicological studies, and considering the potential of C. riparius larvae as a bio-monitoring species, little is known about its genome sequences. This study reports the results of an Expressed Sequence Tags (ESTs) sequencing project conducted on C. riparius larvae using 454 pyrosequencing. METHODS: To gain a better understanding of C. riparius transcriptome, we generated ESTs database of C. ripairus using pyrosequencing method. RESULTS: Sequencing runs, using normalized cDNA collections from fourth instar larvae, yielded 20,020 expressed sequence tags, which were assembled into 8,565 contigs and 11,455 singletons. Sequence analysis was performed by BlastX search against the National Center for Biotechnology Information (NCBI) nucleotide (nr) and uniprot protein database. Based on the gene ontology classifications, 24% (E-value ≤1(-5)) of the sequences had known gene functions, 24% had unknown functions and 52% of sequences did not match any known sequences in the existing database. Sequence comparison revealed 81% of the genes have homologous genes among other insects belonging to the order Diptera providing tools for comparative genome analyses. Targeted searches using these annotations identified genes associated with essential metabolic pathways, signaling pathways, detoxification of toxic metabolites and stress response genes of ecotoxicological interest. CONCLUSIONS: The results obtained from this study would eventually make ecotoxicogenomics possible in a truly environmentally relevant species, such as, C. riparius.