MPRA of Ciona robusta late gastrula whole embryos, RNA Enhancer Barcode, Replicate 3
50 g of the MPRA library was electroporated into ~5000 fertilized eggs. Embryos developed until 5h30 at 22C. Embryos were put into TriZol, and RNA was extracted following the manufacturers instructions (Life Technologies). The RNA was DNase treated using Turbo DnaseI from Ambion following standard instructions. Poly-A selection was used to obtain only mRNA using poly-A biotinylated beads as per instructions (Dyna-beads Life technologies). The mRNA was used in an RT reaction that was specifically selected for the barcoded mRNA (Transcriptor High Fidelity Roche). The RT product was PCR amplified and size selected using Agencourt AMPure beads (Beckman Coulter), then checked for quality and size on the 2100 Bioanalyzer (Agilent) and sent for sequencing on the NovaSeq S4 PE100 mode (Illumina).