2024-11-01
ENA first public
ERC000011
ENA-CHECKLIST
SAMEA114638551
External Id
Dementia Research Institute at KCL, London, UK
INSDC center name
2023-11-17T17:59:36Z
INSDC last update
public
INSDC status
ERS17026689
SRA accession
E-MTAB-13545:FXR1-WT
Submitter Id
ArrayExpress
broker name
HeLa
cell_line
epithelial cell
cell_type
not collected
collection date
human
common name
Protocols: HeLa were grown in 10cm dishes to ~70% confluency and then transfected with either mGFP-FXR1-WT or mGFP-FXR1-V361Pmut for 20 hours. Cells were irradiated once with 150 mJ/cm2 in a Spectroline UV Crosslinker at 254 nm. Cell pellets were snap-frozen 0.1 Units of RNaseI and 4 Units Turbo DNase were added per 1 ml of cell lysate at 1mg/ml protein concentration for RNA fragmentation. Antibodies against GFP (ab290) coupled to magnetic Protein G beads were used to isolate Protein-RNA complexes, and RNA was ligated to a pre-adenylated infra-red labelled IRL3 adaptor. The complexes were then size-separated by SDS-PAGE, blotted onto nitrocellulose and visualised by iBright scanning. RNA was released from membrane by proteinase K digestion and recovered by precipitation. cDNA was synthesized with Superscript IV Reverse Transcriptase (Life Technologies) and AMPure XP beads purification (Beckman- Coulter, USA), then circularised using Circligase II (Epicentre) followed by AMPure XP beads purification. After PCR amplification, libraries were size-selected with Ampure beads and quality controlled for sequencing.
description
adult
developmental stage
adenocarcinoma
disease
WT
genetic modification
endogenous FX1 knock down and transfection with mGFP-FXR1-WT
genotype
not collected
geographic location (country and/or sea)
not applicable
isolate
Homo sapiens
organism
Homo sapiens
organism
cervix
organism part
Homo sapiens
scientific_name
female
sex
FXR1-WT
title