*2.7. Riboprobes Synthesis*

mRNA probes to identify NUCB2B were synthesized by *in vitro* transcription (IVT). Oligonucleotide primers were designed using Primer3 software [45]. Reverse primer contained a T7-promotor sequence to allow direct IVT and the experiment was carried out by means of a DIG RNA labeling mix containing digoxigenin-11-dUTP (Roche, cat. 11277073910). Primers sequences that were used for this study are summarized in Table S2. Primers were diluted to a final concentration of 10 pM. A standard PCR was run to amplify the target region prior to IVT and the amplicon was checked by agarose electrophoresis. Alignment of the obtained sequences was performed by MEGA X software (Molecular Evolutionary Genetics Analysis from www.megasoftware.net) to validate the expected sequence of the amplicon. The concentration of the mRNA probe was measured using the Nanodrop® (Thermo Scientific, Waltham, MA, USA) system.
