Gene discovery through ORESTES sequencing in Triatoma infestans

NICOSIA, S; TEKIEL, V.; BUA, J.; SÁNCHEZ DO.

Carlos Paz, Córdoba, Argentina

Congreso; 44th Annual Meeting – Argentine Society for Biochemistry and Molecular Biology Research (SAIB).; 2008

Argentine Society for Biochemistry and Molecular Biology Research (SAIB).

Triatoma infestans is one of the major vectors of Chagas disease in Argentina. The main objective of this project is to provide new data for exploring alternative strategies towards the development of novel and effective ways to prevent the disease transmission. Expressed Sequence Tags (ESTs) sequencing is a cost-effective and useful advance in gene discovery of human disease vectors, which might result in the identification of new targets for the aforementioned aim.Here, we used the ORESTES (Open Reading frame ESTs) strategy to generate ESTs from mRNA isolated from stage IV nymphs. This strategy preferentially generates ESTs of the central, and thus most informative portion of the transcript, and frequently identifies lessabundant mRNAs. We generated four mini-libraries with the use of arbitrary primers and a low-stringency RT-PCR protocol. From these libraries we obtained 156 sequences that were compared against the NCBI nr database using the BLAST suit of programs. 10% matched T. infestans or other triatomine sequences, 25% were homologous to other organisms, and 26% did not hit any sequence in the database and thus might represent T. infestans-specific sequences. Up to now, we have identify 30 putative new genes for T.infestans, which are represented by those ESTs with positive matches against non T. infestans transcripts.