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Sample GSM6339376

Query DataSets for GSM6339376

Status

Public on May 24, 2023

Title

NDV-infetion HD11 cells rep2

Sample type

SRA

Source name

HD11 cells

Organism

Gallus gallus

Characteristics

cell line: HD11 cells
cell type: Macrophages
genotype: NDV infection

Growth protocol

Chicken-origin macrophage HD11, a permanent chicken bone marrow macrophages cell line was stored in our laboratory and cultured in Dulbecco’s modified essential medium (DMEM) (Solarbio, Beijing, China) containing 10% fetal bovine serum (FBS) (Biological Industries, CT, USA) at 37℃ with 5% CO2.

Extracted molecule

total RNA

Extraction protocol

Total RNA from HD11 cells was extracted at 24 hpi using TRIzol reagent (Invitrogen Corporation, CA, USA) according to the manufacturer’s instruction. RNA concentration and quality of each sample were determined to measure the OD260/OD230 value and OD260/OD280 value by NanoDrop ND-1000 (Thermo Fisher Scientific, Waltham, MA, USA). RNA integrity and gnomic DNA contamination were tested by denatured agarose gel electrophoresis.
MeRIP‑Seq service was performed by Cloudseq Biotech Inc (Shanghai, China). Briefly, RNA from each sample was separately chemically fragmented into fragments with a length of approximately 100 nucleotides using fragmentation buffer (Illumina Inc.). Immunoprecipitation of fragmented RNA was performed with the GenSeq™ m6A-MeRIP Kit (GenSeq Inc., China) according to the the manufacturer’s instructions. Both the input samples without immunoprecipitation and the m6A immunoprecipitation samples were used for RNA-seq library construction with NEBNext® Ultra II Directional RNA Library Prep Kit (New England Biolabs, Inc., USA) following the manufacturer’s instruction. The library quality was evaluated with BioAnalyzer 2100 system (Agilent Technologies, Inc., USA). Library sequencing was subjected to 150 bp paired-end sequencing on an Illumina NovaSeq 6000 instrument.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina NovaSeq 6000

Data processing

Paired-end reads were harvested from Illumina NovaSeq 6000 sequencer, and were quality controlled by Q30. After 3’ adaptor-trimming and low quality reads removing by cutadapt software (v1.9.3). First, clean reads of all libraries were aligned to the reference genome (GCF_000002315.6_GRCg6a ) by Hisat2 software (v2.0.4). Methylated sites on RNAs (peaks) were identified by MACS software. Differentially methylated sites were identified by diffReps. These peaks identified by both softwares overlapping with exons of mRNA were figured out and chosen by home-made scripts. GO and Pathway enrichment analysis were performed by the differentially methylated protein coding genes.
Assembly: GCF_000002315.6_GRCg6a
Supplementary files format and content: The EXCELL table includes the coordinates of the differentially methylated RNA sites in bed format, the length of the methylated RNA sites，the transcript id and gene symbol of differentially methylated RNA，the comparison information calculated by diffreps，and the annotation of the differentially methylated gene for each sample.
Supplementary files format and content: The EXCELL table includes the FPKM value, fold change , p_value, FDR for each sample.

Submission date

Jul 14, 2022

Last update date

May 24, 2023

Contact name

li jin dou

E-mail(s)

ljd18@mails.jlu.edu.cn

Phone

13604365949

Organization name

Jilin University