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Sample GSM3702732 Query DataSets for GSM3702732
Status Public on Aug 12, 2023
Title LCL_input2
Sample type SRA
 
Source name EBV transformed lymphoblastoid cell line
Organism human gammaherpesvirus 4
Characteristics cell type: EBV transformed lymphoblastoid cell lines
passage: 12--15
rip antibody: non
Treatment protocol To reactivate EBV, TPA (20 ng/ml) and Butyric acid (BA, 2.5 mM) were used to treat the cells for 48 hours.
Growth protocol LCL cells were generated in our laboratory. These B-cell lines and PBMC were cultured in RPMI 1640 media (Hyclone, Logan, UT) with 10% BGS (HyClone Bovine Growth Serum).
Extracted molecule total RNA
Extraction protocol The total RNA extraction was performed using Trizol reagent (Invitrogen, Inc., Carlsbad, CA) and treated with DNase I (Invitrogen, Inc., Carlsbad, CA). cDNA was prepared with Superscript II reverse transcriptase kit (Invitrogen, Inc., Carlsbad, CA) according to the manufacturer’s protocol.
The library was prepared using the TruSeq stranded mRNA kit (Illumina, San Diego, CA) according to the manufacturer’s protocol. First, the elute-frag-prime stage was done at 80°C for 2 min to allow annealing without causing fragmentation [12]. RNA was reverse transcribed into first strand cDNA using reverse transcriptase and random primers. This was followed by the second strand cDNA synthesis using DNA Polymerase I and RNase H. The cDNA fragments were used for end repair process with the addition of a single ‘A’ base followed by ligation of adapters. The products were then purified and enriched by PCR amplification for 12 cycles to generate the final RNA-seq library. cDNA libraries were quantified and pooled and subsequent sequencing on an Illumina HiSeq3000 platform 50 bp single read sequencing module.
 
Library strategy RIP-Seq
Library source transcriptomic
Library selection other
Instrument model Illumina HiSeq 3000
 
Description LCL_rpkm
Data processing After sequencing, the first step is quality and adapter trimming. Trimming was conducted by Trim Galore (https://www.bioinformatics.babraham.ac.uk/projects/trim_galore/) to remove adapter sequences and low-quality bases (bases with < 20 quality score will be removed). After trimming, the MeRIP-seq reads were aligned to EBV reference genome by the aligner HISAT2 [39] with the annotation of the splice sites (the EBV reference genome and annotation were downloaded from https://www.ncbi.nlm.nih.gov/nuccore/NC_007605.1). The m6A peaks were called by a graphical model-based peak calling method – MeTPeak [40], with the parameters of window width = 50, sliding step = 50 and read length = 50. The peaks were visualized in IGV (http://www.broadinstitute.org/igv).
Genome_build: EBV genome NC_007605.1
Supplementary_files_format_and_content: peka, rpkm
 
Submission date Apr 02, 2019
Last update date Aug 12, 2023
Contact name Fengchao Lang
E-mail(s) flang@upenn.edu
Organization name University of Pennsylvania
Street address 3610 Hamilton Walk
City PHILADELPHIA
State/province PA
ZIP/Postal code 19104
Country USA
 
Platform ID GPL26377
Series (1)
GSE129217 m6A modification of EBV transcripts
Relations
BioSample SAMN11317995
SRA SRX5624897

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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