 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on May 10, 2016 |
| Title |
Sample siRNA against the MYC gene [1M] |
| Sample type |
SRA |
| |
|
| Source name |
Ascitic fluid of an intestinal-type GC
|
| Organism |
Homo sapiens |
| Characteristics |
sirna: MYC gene
cell line: AGP01
passages: Stomach
|
| Treatment protocol |
An amount of 3x105 cells were seeded into 6cm² plates for each cell line before transfection. Cells were cultured for 24h until cell density was around 50%. A pool or four different double-stranded small interfering RNAs (siRNA) targeting MYC (20 µM, SMARTpool ON-TARGETplus MYC siRNA L-003282-02-0020, Dharmacon, EUA) or scrambled control siRNAs (ON-TARGETplus Non-targeting Pool D-001810-10-05, Dharmacon, GE Lifesciences, EUA) were transfected into AGP01, ACP02 and ACP03 cell lines using Lipofectamine RNAiMAX Transfection Reagent (Life Technologies, EUA). Optimal transfection was reached after 48 h, and total RNA and proteins were extracted with TRIzol reagent. All siRNA experiments were performed three times. In our results, AGP01 scrambled control cells was named 1C, while AGP01 MYC-silenced was called 1M; ACP02 control cells was named 2C, and its MYC-silenced counterpart was represented by 2M; similarly, ACP03 non-targeting control cells was called 3C and the MYC-silenced cells were named 3M.
|
| Growth protocol |
Three GC cell lines previously established and characterized by our group were used: AGP01 (obtained from ascitic fluid of an intestinal-type GC), ACP02 (diffuse-type GC) and ACP03 (intestinal-type GC). Cell lines were cultured in DMEM medium (Gibco/Invitrogen, Germany) supplemented with 10% fetal bovine serum (Gibco/Invitrogen, Germany), 100 U/ml penicilin, 100 µg/ml streptomycin and 0.25 µg/ml amphotericin B. All cultures were maintained in a 5% CO2 air-humidified atmosphere at 37 °C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total mRNA was isolated from gastric cancer cell lines 48h after RNAi transfection using the AllPrep DNA/RNA/Protein Kit (Qiagen, Germany) according to the manufacturer's instructions. Then, mRNA was reverse-transcribed using the High-Capacity cDNA Archive kit according to the manufacturer’s protocol (Life Technologies, USA). The mRNA and cDNA concentration and quality was determined using a NanoDrop spectrophotometer (Kisker, Germany) and 1% agarose gels, respectively. Samples were stored at -80 °C until use.
RNA libraries were prepared for sequencing using standard Ion Torrent protocols
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Ion Torrent PGM |
| |
|
| Data processing |
TMAP to map clean reads to gene reference and/or genome reference
Genes expression level are quantified by a software package: Sailfish.
Analysis of DEG between two samples based on the method of the poisson distribution
Genome_build: hg19
Supplementary_files_format_and_content: tab-delimited text files include RPKM values for each sample obtained by Sailfish tools and DEG screening results based on poisson distribution.
|
| |
|
| Submission date |
May 10, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Daniel Carvalho |
| E-mail(s) |
suporte@genone.com.br
|
| Organization name |
GenOne Biotetechnologies
|
| Street address |
Caixa Postal 26820
|
| City |
Rio de Janeiro |
| State/province |
Rio de Janeiro |
| ZIP/Postal code |
21875-970 |
| Country |
Brazil |
| |
|
| Platform ID |
GPL17301 |
| Series (1) |
| GSE81265 |
Next-Generation Sequencing reveals the effects of MYC silencing in gastric cancer cell lines from northern Brazil |
|
| Relations |
| SRA |
ERX605679 |
| BioSample |
SAMN04966268 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2147867_1M.gene.EXP.txt.gz |
235.6 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Processed data are available on Series record |
| Processed data provided as supplementary file |
| Raw data are available in SRA |
|
|
|
|
 |
