2024-09-30
ENA first public
ERC000037
ENA-CHECKLIST
Biology Centre CAS
INSDC center name
public
INSDC status
ERS21101696
SRA accession
R-P3_2
Submitter Id
22 °C 16hrs (light)/20°C 8 hrs (dark)
air temperature regimen
wild-type
biological status
2017
collection date
thale cress
common name
119100000
estimated size
WT (Col-0)
genotype
Czechia
geographic location (country and/or sea)
missing: lab stock
DD
geographic location (latitude)
missing: lab stock
DD
geographic location (longitude)
growth chamber
growth facility
Seeds were surface sterilised by 70 % and 100 % ethanol series, placed on Petri dishes with cultivation medium and stratified over 2 nights at 4°C, dark. Growth was at LD conditions: 110-120 µmol m-2 s-1, 16hrs light/8 hrs dark, 22 °C during light/20 °C during dark. Seedlisng were grown for 3 days (63-72 hrs) before harvest. Separated shoots and roots were collected by seedling dissection, and the same seedling pool was used as the corresponding shoot and root sample for RNA-seq and ChIP-seq. Replicates 1, 2, and 3 were harvested at zeitgeber time (ZT) 4 – 7, ZT 7 – 10 and ZT 10 – 13, resp.
isolation and growth condition
long-day conditions (110-120 µmol m-2 s-1, 16hrs light/8 hrs dark, light source: OSRAM FQ 54W/840 HO CONSTANT LUMILUX Cool White)
light regimen
4/2 strength Murashige-Skoog (Duchefa M0222)
mineral nutrient regimen
Arabidopsis thaliana
organism
5.8
pH regimen
6d-seedling
plant developmental stage
cultivation medium: 1/2 Murashige-Skoog + 5 mM MES + 0.8% (w/v) plant agar
plant growth medium
cell
plant structure
control
plant treatment
diploid
ploidy
RNA-seq: sample (tissue pool ~ 200 seedlings) was collected into RNAlater (Thermo, ct. no. AM7020) and flash frozen in liquid nitrogen for RNA extraction. Total RNA was extracted using MagMAX™ Plant RNA Isolation Kit (Thermo, cat.no. A33784). polyA-mRNA was enriched using NEBNext® Poly(A) mRNA Magnetic Isolation Module (NEB, cat. No. E7490) and Illumina sequencing libraries were prepared using NEBNext® Ultra™ II RNA Library Prep Kit for Illumina® (NEB, cat. No. E7770); ChIP-seq: sample (tissue pool ~ 400 seedlings) was immediately crosslinked using 1% formaldehyde for 10 min under vacuum,  terminated using 0.125 M glycine for 5 min and flash frozen for ChIP. Sheared chromatin (10 cycles, 30 s ON/ 30 s OFF using a Bioruptor® (Diagenode)) was equally divided between input and samples immunoprecipitated with anti-histone H3 (Merck, cat. no. 07-690); anti-H3K27me3 (Merck, cat. no. 07-449) and IgG (Merck, cat. no.  I5006). Immunocomplexes were collected using Dynabeads® Protein A for Immunoprecipitation (Thermo, cat. no. 10001D) and washed 2 x 5 min with low-salt buffer (containing 150 mM NaCl), 2 x 5 min with high-salt buffer (containing 500 mM NaCl) and 1 x 5 min with LiCl-containing buffer. Immunoprecipitated DNA was purified using iPure kit v2 (Diagenode, cat. no. C03010015). Illumina sequencing library preparation was done using NEBNext® Ultra™ II DNA Library Prep Kit for Illumina® NEB, cat. no. E7645S).
sample material processing
Arabidopsis thaliana
scientific_name
WT_3d_root_autotrophic_rep2
title