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Sample GSM5909429 Query DataSets for GSM5909429
Status Public on Jan 11, 2023
Title HUVEC_ANRIL_1
Sample type SRA
 
Source name ANRIL overexpression in HUVEC cell
Organism Homo sapiens
Characteristics cell line background: HUVEC
genotype/variation: ANRIL overexpression
Treatment protocol GAPDH is used as a control gene to evaluate the over-expression effect of ANRIL. cDNA was synthesized according to standard procedures. Bio-Rad S1000 was subjected to reverse transcription-polymerase chain reaction (RT-PCR) with Bestar SYBR green RT-PCR master mixture (DBI Bioscience, Shanghai, China). The concentration of each transcript was then standardized to GAPDH mRNA level through the 2- ΔΔCT method. Graphpad prism software (San Diego, California) was used for paired Student’s t-test comparison.
Growth protocol HUVEC cells (8000, Sciencell) were cultured at 37 ℃ with 5% CO 2 in ECM with 5% fetal bovine serum (FBS), 100 µg/mL streptomycin, 100 U/mL penicillin and 1% growth factors. Plasmid transfection of HUVEC cells was performed using Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA) according to the manufacturer's protocol. Transfected cells were harvested after 24h for RT-qPCR analysis.
Extracted molecule polyA RNA
Extraction protocol Total RNA was extracted with TRIZOL (Ambion). Total RNA was treated with RQ1 DNase (Promega) to remove DNA. The quality and quantity of the purified RNA were determined by measuring the absorbance at 260nm/280nm (A260/A280) using Nanodrop One (Thermo). RNA integrity was further verified by 1.5% agarose gel electrophoresis
For each sample, 1 μg of total RNA was used for RNA-seq library preparation by KAPA Stranded mRNA-Seq Kit for Illumina® Platforms (#KK8544). Polyadenylated mRNAs were purified by VAHTS mRNA capture Beads (N401-01) and fragmented mRNAs were converted into double strand cDNA. Following end repair and A tailing, the DNAs were ligated to Diluted Roche Adaptor (KK8726). Products corresponding to 300- 500bps were amplified, purified, quantified, and stored at -80 ℃ before sequencing. The strand marked with dUTP (the 2nd cDNA strand) was not amplified, allowing strand-specific sequencing.For high-throughput sequencing, the libraries were prepared following the manufacturer's instructions and applied to Illumina Novaseq 6000 system for 150 nt paired-end sequencing
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina NovaSeq 6000
 
Data processing Illumina bcl2fastq software used for basecalling.
Raw reads containing more than 2-N bases were first discarded. Then adaptors and low-quality bases were trimmed from raw sequencing reads using FASTX-Toolkit (Version 0.0.13). The short reads less than 16nt were also dropped
After that, clean reads were aligned to the GRch38 genome by tophat2 (Kim, Pertea et al. 2013) allowing 4 mismatches.
The R Bioconductor package edgeR was utilized to screen out the differentially expressed genes (DEGs). A false discovery rate <0.05 and fold change>2 or < 0.5 were set as the cut-off criteria for identifying DEGs
The ABLas pipeline was applied to define and quantify the alternative splicing events (ASEs) and regulated alternative splicing events (RASEs) between the samples according to previously described methods
Genome_build: GRCh38
Supplementary_files_format_and_content: tab-delimited text files include gene FPKM values for each Sample
 
Submission date Feb 21, 2022
Last update date Jan 11, 2023
Contact name Dong Chen
Organization name ABLife, Inc.
Department Center for Genome Analysis
Street address 388 GaoXin 2nd Road, East Lake Hi-Tech Development Zone
City Wuhan
State/province Hubei
ZIP/Postal code 430075
Country China
 
Platform ID GPL24676
Series (1)
GSE197115 ANRIL overexpression globally induces expression and alternative splicing of genes involved in inflammation in HUVECs
Relations
BioSample SAMN26137385
SRA SRX14240876

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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