2025-07-14
ENA first public
ERC000011
ENA-CHECKLIST
Norwegian University of Life Sciences (NMBU)
INSDC center name
public
INSDC status
ERS25125916
SRA accession
E-MTAB-15297:Pabies_Embryo_G20_R1
Submitter Id
144
hour
age
ArrayExpress
broker name
Clonal line ID#B10W (cross: #26509 × #2707)
clonal line
not collected
collection date
Norway spruce
common name
Protocols: Embryogenic callus cultures of Picea abies (line B10W), derived from a single zygotic embryo, were maintained in vitro on proliferation medium under dark conditions at 23 ± 1°C. Callus aggregates were exposed to continuous gamma radiation from a ⁶⁰Co source at specified dose rates (0–100 mGy/h) for 144 hours. At the end of exposure, callus samples were harvested, flash-frozen in liquid nitrogen, and stored at −80°C prior to RNA extraction. Embryogenic callus cultures of Picea abies (line ID#B10W) were maintained in vitro on proliferation medium (PM) composed of AL basal medium with inositol, modified vitamin mix, 10 μM 2,4-D, 5 μM BA, 1% sucrose, and 0.3% Phytagel. Cultures were grown in 9 cm Petri dishes at 23 °C in darkness prior to gamma irradiation. In vitro callus cultures were exposed to continuous gamma radiation for 144 hours at dose rates of 0 (control), 1, 10, 20, 40, or 100 mGy/h using a ⁶⁰Co source at the FIGARO low dose irradiation facility (NMBU). Petri dishes were positioned at calibrated distances and rotated midway to ensure even exposure. Controls were kept shielded from radiation in the same room. Total RNA was extracted using the MasterPure Complete DNA and RNA Purification Kit (Epicentre) following the manufacturer’s protocol, with modifications: 1–2% polyvinylpyrrolidone (PVP, mw 360,000) was added to the extraction buffer, and 1,4-dithiothreitol (DTT) was replaced with 3 μl beta-mercaptoethanol. RNA was quantified using a NanoDrop ND-1000 and assessed for quality using an Agilent 2100 Bioanalyzer. Strand-specific RNA-seq libraries were prepared using the TruSeq stranded mRNA library preparation protocol (Illumina) according to the manufacturer’s instructions. Libraries were constructed from total RNA and sequenced as 150 bp paired-end reads.
description
wild type genotype
genotype
not collected
geographic location (country and/or sea)
not applicable
isolate
Picea abies
organism
Picea abies
organism
embryo-derived callus
organism part
1
replicate
Picea abies
scientific_name
Pabies_Embryo_G20_R1
title