title: 0dpa_1
organism: Xenopus tropicalis
description: Protocols: Xenopus tropicalis tadpoles at stage NF50 were anaesthetised in 0.1% MS222 in 0.01X Marc's Modified Ringer (MMR) solution (10mM NaCl, 0.2mM KCl, 0.1mM MgSO4, 0.2mM CaCl2, 0.5mM HEPES, pH 7.4) The spinal cords were manually dissected and immediately transferred in Trizol. Total RNAs were isolated from pools of 20 spinal cords using Trizol. After precipitation using 15µg of linear acrylamide as a carrier, the RNAs were resuspended in 50µL DNAse-free water. To remove any residual DNA, 2µL of TURBO DNase I was added to each sample. After a 15min incubation at 37ºC and 5min at 65ºC, the RNAs were purified by phenol-chlorophorm extraction and precipitation. The pellets were washed with 70% EtOH and resuspended in 12µL nuclease-free water. RNA concentrations were determined using Qubit HS RNA assay kit (Invitrogen) on the Qubit Fluorometer 2.0 (Invitrogen) and ranged from 30 to 70ng.µL-1. RNA Integrity was tested with the Agilent RNA 6000 Pico kit on the Agilent Bioanalyser. All samples with the RNA Integrity Number (RIN) ≥7 were regarded as RNA of acceptable quality. Finally, an aliquot of each sample was reverse-transcribed and a PCR with oligonucleotides specific for ODC was performed to confirm the integrity of the RNAs before being process for RNAseq. Enrichment of polyadenylated transcripts with oligo-dT beads; after washing everything else away, mRNA was fragmented and ds-cDNA prepared
attributes.tag: ENA-FIRST-PUBLIC
attributes.value: 2020-02-26T17:08:15Z
attributes.tag: ENA-LAST-UPDATE
attributes.value: 2020-02-11T13:25:13Z
attributes.tag: External Id
attributes.value: SAMEA6534875
attributes.tag: INSDC center name
attributes.value: University of Manchester Faculty of Biology, Medicine and Health School of Medical Sciences Division of Developmental Biology and Medicine
attributes.tag: INSDC first public
attributes.value: 2020-02-26T17:08:15Z
attributes.tag: INSDC last update
attributes.value: 2020-02-11T13:25:13Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-8785:0dpa_1
attributes.tag: age
attributes.value: 3
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: common name
attributes.value: tropical clawed frog
attributes.tag: developmental stage
attributes.value: NF stage 50
attributes.tag: genotype
attributes.value: wild type genotype
attributes.tag: individual
attributes.value: mixed embryo pool
attributes.tag: organism part
attributes.value: spinal cord
attributes.tag: sample name
attributes.value: E-MTAB-8785:0dpa_1
attributes.tag: scientific_name
attributes.value: Xenopus tropicalis
attributes.tag: sex
attributes.value: mixed