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Sample GSM7916786 Query DataSets for GSM7916786
Status Public on Dec 08, 2023
Title SR34a-GFP iCLIP2 ABA biol rep 1
Sample type SRA
Source name 44h germinated seeds + 2h ABA treatment
Organism Arabidopsis thaliana
Characteristics tissue: 44h germinated seeds + 2h ABA treatment
genotype: promSR34a:SR34a-GFP in sr34a-1
treatment: ABA 0.5 µM
Treatment protocol 44h germinated seeds from promSR34a:SR34a-GFP sr34a-1 were transferred to MS medium (Control) or MS medium supplemented with 0.5 µM ABA during 2h. 44h germinated seeds from prom35S:GFP sr34a-1 were transferred to MS medium (Control) or MS medium supplemented with 0.5 µM ABA during 2h and pooled together before iCLIP2.
Growth protocol Stratified seeds were grown on MS medium in continuous white light (100 µmol/m2) at 21 ºC for 44h
Extracted molecule total RNA
Extraction protocol Germinated seed tissues were UV-crosslinked (2000 mJ/cm2), frozen in liquid N2 and ground to a homogeneous powder before cell lysis. SR34a-GFP-RNA and GFP-RNA complexes were precipitated using GFP-trap agarose beads (Chromotek). After 3' end RNA linker ligation and 5'end radiolabelling, RNA-protein complexes were run on an SDS gel, transferred on a nitrocellulose membrane, and revealed by autoradiography. After a proteinase K treatment, protein-bound RNA was purified using phenol-chloroform and concentrated with the RNA Clean and Concentrator kit (Zymo Research, R1013).
Library preparation was performed exactly as in Buchbender et al., 2019 (doi:10.1016/j.ymeth.2019.10.003). Purified RNAs were reverse transcribed and 5' adaptors were ligated to the subsequent cDNAs. cDNAs were then PCR-amplified. 10 nM of each library was submitted to sequencing using the NextSeq 500 system (Illumina).
Library strategy OTHER
Library source transcriptomic
Library selection other
Instrument model Illumina NextSeq 500
Data processing Reads were demultiplexed and sequencing adapters removed using Flexbar (3.5.0). Reads were then quality- and length-trimmed with Flexbar. A genome index was created using STAR (2.7.3a) with the Arabidopsis thaliana genome version TAIR10 and gene annotation from Araport (version 11). Reads were then mapped using STAR and the created genome index, allowing only softclipping of 3’ ends (--alignEndsType Extend5pOfRead1). PCR duplicates were removed using umi_tools (1.1.4). The uniquely mapped and deduplicated reads from each replicate were merged and peak called with PureCLIP (1.3.1) in standard mode.
Assembly: TAIR10
Supplementary files format and content: Bedgraph files for visualisation of iCLIP2 peaks using a genome viewer software
Library strategy: iCLIP2
Submission date Nov 24, 2023
Last update date Dec 08, 2023
Contact name Tom Laloum
Phone 918995207
Organization name Instituto Gulbenkian de Ciencia
Lab Plant Molecular Biology
Street address Rua da Quinta Grande, 6
City Oeiras
ZIP/Postal code 2780-156
Country Portugal
Platform ID GPL19580
Series (2)
GSE248560 Identification of SR34a direct RNA targets in germinated seeds grown for 44 hours and exposed or not to ABA for 2 hours
GSE248563 sr34a-1 mutant germinated seeds and exposure to ABA
BioSample SAMN38412747
SRA SRX22635858

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA

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