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| Status |
Public on May 10, 2019 |
| Title |
H_control |
| Sample type |
SRA |
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| Source name |
H1299
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| Organism |
Homo sapiens |
| Characteristics |
genotype/variation: sgRNA control
cell line: H1299
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| Growth protocol |
H1299 cells were cultured in Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin (Invitrogen).
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from cell lines by NucleoSpin RNA XS (Macherey Nagel, Duren, Germany).
A total amount of 3 μg RNA per sample was used as in put material for the RNA sample preparations. Sequencing libraries were generated using NEBNext® Ultra™ RNA Library Prep Kit for Illumina® (NEB, USA) following manufacturer’s recommendations and index codes were added to attribute sequences to each sample. Briefly, mRNA was purified from total RNA using poly-T oligo-attached magnetic beads. Fragmentation was carried out. using divalent cations under elevated temperature in NEBNext First Strand Synthesis Reaction Buffer（5X）. First strand cDNA was synthesized using random hexamer primer and M-MuLV Reverse Transcriptase（RNase H. Second strandcDNA synthesis was subsequently performed using DNA Polymerase I and RNase H. Remaining overhangs were converted into blunt ends via exonuclease/polymerase activities. After adenylation of 3’ ends of DNA fragments, NEBNext Adaptor with hairpin loop structure were ligated to prepare for hybridization. In order to select cDNA fragments of preferentially 150~200 bp in length, the library fragments were purified with AMPure XP system (Beckman Coulter, Beverly, USA). Then 3 μl USER Enzyme (NEB, USA) was used with size-selected, adaptor-ligated cDNA at 37°C for 15 min followed by 5 min at 95 °C before PCR. Then PCR was performed with Phusion High-Fidelity DNA polymerase, Universal PCR primers and Index (X) Primer. At last, PCR products were purified (AMPure XP system) and library quality was assessed on the Agilent Bioanalyzer 2100 system.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
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| Data processing |
The RNA-seq reads of knockdown groups were first processed with Trimmomatic (Trimmomatic-0.38) to eliminate the adapter contamination.
The qualified reads from knockdown groups were aligned to the reference genome by STAR (STAR_2.6.1d).
The qualified reads of knockdown groups were conducted with RSEM (RSEM-1.3.1). Gene expression quantification were conducted with FPKM (Fragments Per Kilobase of transcript per Million mapped reads).
The reference genome and gene annotations were retrieved from Ensembl database.
Genome_build: GRCh38.p12
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample
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| Submission date |
Apr 10, 2019 |
| Last update date |
May 10, 2019 |
| Contact name |
JOU-HO SHIH |
| E-mail(s) |
roromanworld@gmail.com
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| Organization name |
Institute of Biomedical Sciences, Academia Sinica
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| Street address |
128 Sec. 2, Academia Rd. Nankang, Taipei
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| City |
Taipei |
| State/province |
Please Select |
| ZIP/Postal code |
11529 |
| Country |
Taiwan |
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| Platform ID |
GPL24676 |
| Series (1) |
| GSE114826 |
RNA sequencing results of wild type and overexpresssion or knockdown PTTG3P in LUAD cell lines based on IlluminaNovaSeq 6000 platform |
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| Relations |
| BioSample |
SAMN11382776 |
| SRA |
SRX5661291 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3716410_H_control.Gene.fpkm.txt.gz |
209.1 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Processed data provided as supplementary file |
| Raw data are available in SRA |
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