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Sample GSM3972548

Query DataSets for GSM3972548

Status

Public on May 18, 2021

Title

Kidney Control Rep1

Sample type

SRA

Source name

kidney

Organism

Rattus norvegicus

Characteristics

strain: Sprague-Dawley
tissue: kidney
age: three-month-old
diagnosis: control

Treatment protocol

Briefly, rats were deprived of water for 48 hours and then anaesthetized by pentobarbital sodium (40 mg/kg, ip.). Catheters were palced in the right femoral vein and the common carotid artery (24 G×21 mm, SPECATH, Foshan, China), and a baseline arterial blood sample was drawn (1 ml) for determination of SCr. Then, INDO was administreted intravenously (10 mg/kg, iv.), and followed after 15 minuters by L-NAME (10 mg/kg, iv.). Another 15 minuters later, rats were randomized to receive iopromide (CI-AKI group) or normal saline (control group) via the carotid artery cannulation (7.8 ml/kg, ia.). The rats were then allowed to recover in individual cages with free access to tap water and standard chow. Approximately 12 hours after iopromide injection, the blood sample for the measurement of postoperative SCr was obtained under anesthesia. The kidney tissue for RNA-sequencing or qRT-PCR validation was then harvested immediately and stored in the RNAsafty Reagent (Bohao, Shanghai, China) at 4 degrees Celsius overnight, before tansfered to an minus 20 degrees Celsius refrigerator. Part of the kidney tissue, containing cortex and medulla, were fixed in 10 % neutral formalin liquid for histological analysis. The sections of kidney tissue were stained with hematoxylin and eosin, and the Paller scores were calculated to determind the severity of the tubular injury. In the end, the rats were sacrificed with an overdose of pentobarbital anesthesia (200 mg/kg, ip.).

Growth protocol

The three-month-old male Sprague-Dawley rats (Fujian Medical University breed, China), weighing approximately 250 g at the start of the experiment, were used and kept in individual cages under controlled conditions of light (12 hours/12 hours light/dark cycle) and temperature (21~23℃), with free access to tap water and standard rat chow for a 7-day adaptive period.

Extracted molecule

total RNA

Extraction protocol

Total RNA was extracted using mirVanaTM miRNA Isolation Kit (Cat# AM1561, Ambion)following the manufacturer’s instructions and checked for a RIN number to inspect RNA integrity by an Agilent Bioanalyzer 2100 (Agilent technologies, Santa Clara, CA, US).Qualified total RNA was further purified by RNAClean XP Kit (Cat A63987, Beckman Coulter,Inc.Kraemer Boulevard Brea, CA,USA)and RNase-Free DNase Set (Cat#79254, QIAGEN, GmBH, Germany).
RNA libraries were prepared for sequencing using standard Illumina protocols

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

HiSeq X Ten

Data processing

Sequenced raw reads were preprocessed by removing rRNA reads, sequencing adapters, short-fragment reads and other low-quality reads with Seqtk (https:// github.com/lh3/seqtk).
Obtained clean reads were mapped to the Rat rn6 reference genome with two mismatches by Hisat2 (version 2.0.4). After genome mapping, Cufflinks v2.1.1 was run with a reference annotation to generate FPKM (fragments per kilobase of exon model per million mapped reads)values for known gene models.
The p-value significance threshold was set by the false discovery rate (FDR) strictly controlled using the Benjamini and Hochberg algorithm. The fold-changes were estimated according to the FPKM in each sample. Differentially expressed genes were identified using Cuffdiff with the following filter criteria: FDR≤0.05 and fold-change≥2.
Genome_build: rn6

Submission date

Jul 25, 2019

Last update date

May 18, 2021

Contact name

Zhiqing Wang

E-mail(s)

wzq137747@163.com

Phone

+8618905911812

Organization name

900 Hospital of the Joint Logistics Team