Small RNA sequencing from Xenopus laevis egg
To characterise the population of miRNA during Xenopus embryo development, different developmental stages were considered: unfertilised eggs, and developmental embryos at stages 7, 11-12, 18-20, 34-36 and 56. Total RNA was isolated using TRIzol® (15596-026, Life technologies, UK) according to manufacturer´s recommendations and DNase treated. RNA integrity and purity were assessed in 1% (w/v) agarose gel stained with ethidium bromide and on a Nanodrop ND-1000 (Thermo Scientific, DE, USA), respectively. Ratios of Abs 260/280 and Abs 260/230 were above 2.0 for all RNA   The sRNA libraries were prepared as described before using total RNA from the different developmental stages. Briefly, the RNA was ligated to the adaptors, reverse transcribed, PCR amplified and purified from a polyacrylamide gel. cDNA libraries were sequenced using an Illumina Genome Analyzer II.