Sequencing of Bothrops jararaca genome to characterize toxins gene structure and their regulatory elements

Abstract

Most studies involving snake venoms were based on traditional approaches like Biochemistry and Molecular Biology, contributing to many fields of biological sciences. The global characterization techniques, such as transcriptomics and proteomics, enriched this knowledge by indicating the complex gene organization of toxin families and the presence of important molecular evolutionary processes, such as accelerated evolution, exon shuffling, functional convergence, retrotransposition, etc. Surprisingly, despite all acquired knowledge, the molecular basis of snake genomes is poorly known. Even in the Era of Next Generation Sequencing (NGS), no genome of poisonous snake, Brazilian or not, has been resolved. Furthermore, the "Squamata" (snakes and lizards) group remains unexplored particularly when compared to many vertebrate groups.In this project, we propose to achieve a high coverage of the genome sequence of Bothrops jararaca - the major medically important species in Brazil. To do so, we will build shotgun and mate-pair libraries to perform Next Generation Sequencing (NGS) using Illumina technology. The mate-pair libraries will be constructed using fragments of different sizes to provide a better mapping, as it was shown to achieve better results for the resolution of other vertebrates genomes. Longer sequences (>400bp) will be obtained by the 454 technology with the Roche-GSFLX equipment. BAC libraries will also be constructed and sequenced in 454-GSJunior equipment in order to obtain continuous and longer segments of the genome. These BACs could be randomly selected or screened with probes consisted of cDNAs from toxins of interest. All generated sequences will be assembled in contigs and scaffolds of the genome. These will allow us to obtain a large collection of genome segments that may be selected in silico. Therefore, we hope to identify and characterize genes of toxins, genes involved in venom production and secretion, retrotransposable elements and other elements with relevance in the evolution of the venom system. (AU)