|
| Status |
Public on Feb 13, 2019 |
| Title |
NP_HuR knockdown 3 |
| Sample type |
SRA |
| |
|
| Source name |
Rat nucleus pulposus_HuR knockdown
|
| Organism |
Rattus norvegicus |
| Characteristics |
strain background: Wistar
cell type: primary rat nucleus pulposus (NP) cells
genotype/variation: shHuR-transduced
culture condition: hypoxia (1% O2)
|
| Treatment protocol |
Rat NP cells were transduced with lentivirally-delivered control (MISSION PLKO.1 non-target ShRNA, ShCTR) or HuR-specific (ShHuR) shRNA. 24 h before collection of total mRNA, cells were cultured under 1% O2 conditions.
|
| Growth protocol |
Primary rat nucleus pulposus (NP) cells were dissected from Wistar rat spines, expanded from explant tissues, and maintained in DMEM containing 10% FBS and antibiotics
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total mRNA was extracted using Qiagen RNeasy mini columns and treated with DNAse1
Illumina TruSeq Stranded Total RNA Kit with Ribo-Zero GoldTotal RNA samples (100-500 ng) were hybridized with Ribo-Zero Gold to substantially deplete cytoplasmic and mitochondrial rRNA from the samples. Stranded RNA sequencing libraries were prepared as described using the Illumina TruSeq Stranded Total RNA Kit with Ribo-Zero Gold (RS-122-2301 and RS-122-2302). Purified libraries were qualified on an Agilent Technologies 2200 TapeStation using a D1000 ScreenTape assay (cat# 5067-5582 and 5067-5583). The molarity of adapter-modified molecules was defined by quantitative PCR using the Kapa Biosystems Kapa Library Quant Kit (cat#KK4824). Individual libraries were normalized to 10 nM and equal volumes were pooled in preparation for Illumina sequence analysis.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
ShHuR 3
13657X12_S3_shHuR_Hx_normalized
|
| Data processing |
HiSeq 50 Cycle Single-Read Sequencing version 4Sequencing libraries (25 pM) were chemically denatured and applied to an Illumina HiSeq v4 single read flow cell using an Illumina cBot. Hybridized molecules were clonally amplified and annealed to sequencing primers with reagents from an Illumina HiSeq SR Cluster Kit v4-cBot (GD-401-4001). Following transfer of the flowcell to an Illumina HiSeq 2500 instrument (HCSv2.2.38 and RTA v1.18.61), a 50 cycle single-read sequence run was performed using HiSeq SBS Kit v4 sequencing reagents.
Genome_build: Rn5 Ensembl annotations (Build 75)
Supplementary_files_format_and_content: File containing Gene identifiers, log2FC, adusted p value, and normalized reads
|
| |
|
| Submission date |
Feb 19, 2018 |
| Last update date |
Feb 13, 2019 |
| Contact name |
Makarand Risbud |
| E-mail(s) |
makarand.risbud@jefferson.edu
|
| Phone |
(215) 955-1063
|
| Organization name |
Thomas Jefferson University
|
| Department |
Orthopaedic Surgery
|
| Lab |
511 College Bldg.
|
| Street address |
1015 Walnut St.
|
| City |
Philadelphia |
| State/province |
PA |
| ZIP/Postal code |
19107 |
| Country |
USA |
| |
|
| Platform ID |
GPL18694 |
| Series (1) |
| GSE110804 |
RNA sequencing reveals RNA binding protein HuR/ELAVL1 regulates extracellular matrix gene expression in nucleus pulposus cells of the intervertebral disc |
|
| Relations |
| BioSample |
SAMN08564449 |
| SRA |
SRX3721472 |
