Parallel DNA pyrosequencing unveils new zebrafish microRNAs

Abstract

Background: MicroRNAs (miRNAs) are a new class of small RNAs of approximately 22 nucleotides in length that control eukaryotic gene expression by fine tuning mRNA translation. They regulate a wide variety of biological processes, namely developmental timing, cell differentiation, cell proliferation, immune response and infection. For this reason, their identification is essential to understand eukaryotic biology. Their small size, low abundance and high instability complicated early identification, however cloning/Sanger sequencing and new generation genome sequencing approaches overcame most technical hurdles and are being used for rapid miRNA identification in many eukaryotes.

A bioinformatics-based update on microRNAs and their targets in rainbow trout (Oncorhynchus mykiss)

“…MiRNAs can be identified by direct cloning (Ramachandra et al, 2008;Sunkar et al, 2005), high-throughput sequencing (Chi et al, 2011;Soares et al, 2009) and computational homology-based methods (Jones-Rhoades and Bartel, 2004;Li et al, 2010). And recently, a lot of miRNAs have been identified by high-throughput sequencing in various animals and plants for its high performance, including rainbow trout (Ma et al, 2012).…”

“…Interestingly, based on dre-miR-2189, we have identified 11 mir 2189 members in rainbow trout. However, there is currently only one mir-2189 from zebrafish in the miRBase, which has been detected by deep sequencing (Soares et al, 2009). It suggests that O. mykiss has more dependence on this family.…”

Comprehensive identification and profiling of Nile tilapia (Oreochromis niloticus) microRNAs response to Streptococcus agalactiae infection through high-throughput sequencing

“…The SOLiD™ system has emerged as a powerful tool for the large-scale identification of miRNAs and the analysis of their expression especially for those species with limited genomic information [5,16]. These species include shrimp [17], zebrafish [18], medaka [19], atlantic halibut [20], common crap [21], catfish [22], Japanese flounder [5], grouper [6] and Nile tilapia [14].…”

Computational Tools for Identification of microRNAs in Deep Sequencing Data Sets

“…For this, cDNA libraries are prepared following commonly used protocols (Droege M and Hill B. 2008;Soares et al 2009). These libraries contain specific adaptors for the small RNA molecules containing specific priming sites for sequencing.…”

“…In other words, miRNAs play an important role in zebrafish organogenesis and their expression at specific time points is relevant to organ formation and differentiation. Since identification of the complete set of miRNAs is fundamental to fully understand biological processes, we have used high throughput 454 DNA pyrosequencing technologies to fully characterize the zebrafish miRNA population (Soares et al 2009). For this, a series of cDNA libraries were prepared from miRNAs isolated at different embryonic time points and from fully developed organs sequenced using the Genome Sequencer TM FLX.…”