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| Status |
Public on Dec 15, 2020 |
| Title |
control-3 |
| Sample type |
SRA |
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| Source name |
OE19 oesophageal adenocarcinoma cell line
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| Organism |
Homo sapiens |
| Characteristics |
cell type: oesophageal adenocarcinoma derived cell line
treatment: miRCURY LNA Power microRNA Inhibitor Negative Control A (Qiagen, #EX-199006-101)
cell line: OE19
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| Treatment protocol |
24 h after seeding, cells at 80 % confluence were irradiated with a dose of 2 Gy, chosen in concordance with previously reported irradiation protocols [6, 9], in an X-Rad 320 irradiation machine (Precision X-Ray, North Branford, CT, USA). Negative controls were mock-irradiated. Samples were irradiated in full scatter conditions with 2 Gy at a SSD of 65 cm with HVL of 2.00 mm Aluminium, at 300 kVp with the tube current at 13 mA at a dose-rate of 2.3 Gy/min. The accuracy of the dose-measurement was ± 5%. The dose calibration of the orthovoltage 300 kVp X-ray beam produced by the X-RAD 320 was performed according to the Institute of Physics and Engineering in Medicine and Biology (IPEMB) protocol.
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| Growth protocol |
The OE19 cell line (purchased from Sigma ECACC ) was cultured using RPMI 1640 medium (Thermo Fisher Scientific, Scoresby, Australia) supplemented with 10 % foetal bovine serum (FBS) (Thermo Fisher Scientific, #10099141, Scoresby, Australia), 50 U/mL penicillin (Thermo Fisher Scientific, #15070063, Scoresby, Australia), 50 µg/mL streptomycin (Thermo Fisher Scientific, #15070063, Scoresby, Australia), and 100 µg/mL normocin (InvivoGen, #nta-nr-2, San Diego, USA) and cultured using standard techniques and reagents
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA isolation, including DNase digestion, was performed using the miRNeasy Mini Kit (Qiagen, #217004, Chadstone, Australia) and RNase-free DNase Set (Qiagen, #79254, Chadstone, Australia) as instructed by the manufacturer. Quantification of the final RNA concentration was performed via UV spectrophotometry (NanoDrop™ 2000 Spectrophotometer, Thermo Fisher Scientific, Wilmington, USA).
Libraries of small RNAs were prepared from 150 ng of RNA using the NEBNext™ kit (New England Biolabs). After reverse transcription, 15 cycles of PCR were performed to enrich for successfully ligated molecules. The libraries were size selected using 3 % agarose gels targeting a 122–182 bp range. Size-selected libraries were run on Bioanalyzer high-sensitivity DNA chips to confirm size, minimal adapter dimers, and to estimate yield.
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| Library strategy |
ncRNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina NovaSeq 6000 |
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| Data processing |
Raw reads were trimmed and filtered for short sequences using cutadapt v.1.8 [105]. Trimmed FASTQ files, averaging 20 million reads per sample, were quality checked using the FastQC program (v.0.11.8)
Reads were mapped against the human reference genome (Gencode GRCh38) using STAR aligner (version:2.7.0f_0328)
RNA counts were within-lane normalised for differences in GC content and Length using the EDASeq package in R [108], and then between-lane normalized for differences in read depth using the DESeq 2 package in R
Differential expression was assessed via Mann Whitney U test in R.
Genome_build: Gencode GRCh38
Supplementary_files_format_and_content: The mRNA NGS processed data file contains a matrix of normalised data for the samples vs. mRNAs
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| Submission date |
Nov 10, 2020 |
| Last update date |
Dec 15, 2020 |
| Contact name |
George Mayne |
| E-mail(s) |
george.mayne@flinders.edu.au
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| Phone |
+61 08 8204 6088
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| Organization name |
Flinders University of South Australia
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| Department |
Surgery
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| Lab |
3d213
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| Street address |
Room 3D213, Dept of Surgery, Flinders Medical Center
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| City |
Bedford Park |
| State/province |
SA |
| ZIP/Postal code |
5042 |
| Country |
Australia |
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| Platform ID |
GPL24676 |
| Series (2) |
| GSE161148 |
MicroRNA profiling in oesophageal adenocarcinoma cell lines and patient serum samples reveals a role for miR-451a in radiation resistance [control and miR-451a inhibitor treatments] |
| GSE161149 |
MicroRNA profiling in oesophageal adenocarcinoma cell lines and patient serum samples reveals a role for miR-451a in radiation resistance |
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| Relations |
| BioSample |
SAMN16714486 |
| SRA |
SRX9467028 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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