• DGE
• EIRA
• Neural Induction
• RNA dynamics
• Transcriptomics

Dynamic stability of messenger RNAs (mRNAs) is of central importance in the development and maintenance of an organism. Exon-Intron Ratio Analysis (EIRA) enables assessment of the dynamic stability of mRNAs during development. The trend of the ratio between the number of exonic and intronic reads (EIR, Exon-Intron Ratio) in a transcriptome obtained by Deep Sequencing will be tracked. The value of EIR is used as a proxy for the stability of a given messenger. Factors that may affect the EIR value include the action of microRNAs (miRNAs), short RNA sequences capable of recognizing and pairing with the 3' UTR region of mRNAs. Their binding leads to destabilization of the messenger itself through a degradation process or interference with translation, thus contributing to post-transcriptional control of translation.
Xenopus laevis is a widely used model system in many evolutionary and developmental studies, offering numerous experimental advantages. Small portions of tissue belonging to the animal pole, Animal Caps (ACs), constitute the early neural induction model used in many and the present study. ACs were taken from control embryos, and embryos injected with noggin neural inducer mRNA. ACs were harvested and grown to different stages of embryonic development (9, 10.5, 14 and 22). Bioinformatic analysis on the transcripts made it possible to calculate the EIR value for each sequence, at each stage, and for all replicates. Using clustering methods, groups of sequences whose EIR value varied significantly between control and Noggin treatment were identified. Given the importance of miRNAs in interference phenomena on mRNAs, the effect, in terms of neuralization, of miRNA suppression was evaluated in a pilot experiment. This is accomplished by knocking down the function of Dicer1, a nuclease involved in the process of miRNA maturation. To evaluate the effects on neuralization produced by Dicer1 knockdown, preliminary RT-PCRs targeting neural marker genes were performed.