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Clonación de genes por spliced leader a partir de genotecas de expresión de cisticercos de Taenia solium

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Author(s): Oswgladys Garrido | Dayana Requena | Carlos Flores Angulo | Teresa Gárate | Elizabeth Ferrer

Journal: Salus Online
ISSN 1316-7138

Volume: 16;
Issue: 1;
Start page: 13;
Date: 2012;
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Keywords: Taenia solium | cysticercosis | Spliced Leader | cloning.

ABSTRACT
Cysticercosis is caused by the larval stage of Taenia solium (cysticercus). The diagnosis of the disease is limited by the availability of parasite antigens; an alternative would be the cloning of gene encoding antigens. In T. solium, as in other parasites, an alternative mechanism in the processing of some mRNAs called transsplicing occurs, in which a small RNA known as Spliced Leader (SL) is added to the 5´ end of pre-mRNA molecules, forming a common 5´-terminal exon of the mature mRNAs. Due to limitations for diagnosing the disease, in addition to the interest in the study of this mechanism, the aim of this work was to clone molecules that use this post-transcriptional processing. In this study we did a screening by PCR from cDNA library of T. solium cysticerci using the forward primer TSSL-DW2 and the reverse primer ZAP-3´UP that hybridize with SL and vector sequence, respectively. cDNAs of different sizes were obtained that were cloned in maintenance plasmids (pGEM-Teasy). The presence of inserts and their sizes were estimated by colony PCR, obtaining a total of 56 clones of different sizes (500-1200 bp). This design allows the identification of of T. solium genes using the trans-splicing mechanism; and besides being an easy strategy to clone complete molecules, it opens the way for future investigations on the diagnosis of cysticercosis
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