|
| Status |
Public on Aug 25, 2015 |
| Title |
High grade tumor sample [Ast_15Agosto2014_3_(HTA-2_0)] gene-level |
| Sample type |
RNA |
| |
|
| Source name |
Tumor sample
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: glia tissue
tumor grade: GBM
|
| Treatment protocol |
A total of eight astrocytic tumors and two control tissues were collected during the period of 2012–2014: four low-grade Ast (three pilocytic Ast (PAst) and one diffuse Ast (DAst)) and four high-grade Ast (four glioblastoma multiforme (GBM)). Brain tumor samples were collected at surgery, snap frozen in liquid nitrogen and stored at −80 °C until use. All tumors were collected from children – one through 15 years of age – refer from the Children´s hospital, National Medical Center Century XXI, Mexican Institute of Social Security. All patients were from families with no history of cancer (de novo tumors)Control samples were obtained from pediatric patients (one boy and one girl) who had submitted to temporal lobe resection for epilepsy. Tumors were histopathologically classified by two pathologists according to a criteria established by the WHO.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted from tissue samples using TRIzol (Ambion life technologies, Thermo Scientific Inc.) following the methodology described by Hongbao et al. 2008 and with certain modifications described by Vujovic et al. 2013. Briefly, tumor samples were homogenized in 1 ml of TRIZOL® Reagent (1ml per 50 mg of tissue), and incubated at room temperature (25°C) for 5 min. After that, we added 200 µl of chloroform (per ml TRIzol) and shake vigorously the samples for 15 sec. Then, samples were incubating at 4°C for 15 min and they were centrifuged at 12,000xg for 15 min (4°C). The aqueous phase, which corresponds to RNA was transferred to a new 1.5 ml tube and RNA was precipitated adding 500 µl isopropanol (per ml TRIzol) and incubating for 30 min at room temperature. After that, samples were centrifuged at 12,000xg (10 min) at 4°C, the supernatant was removed and pellet was washed once with 75% ethanol (1 ml/ml TRIzol), vortexed and centrifuged at 7,500xg for 5 min at 4°C. The final pellet was dissolved in 300-600 µl of RNase-free water and it was stored at -80 °C until use.
|
| Label |
biotin
|
| Label protocol |
Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 6 ug total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).
|
| |
|
| Hybridization protocol |
Following fragmentation, 10 ug of cRNA were hybridized for 16 hr at 45C on Affymetrix HTA 2.0 Array. Chips were washed and stained in the Affymetrix Fluidics Station 400.
|
| Scan protocol |
GeneChips were scanned using GeneArray Scanner G2500A.
|
| Description |
Gene expression data from pediatric brain tumor grade V
|
| Data processing |
For data analysis two softwares were used: Expression Console and Transcript Analysis Console. The first software allows us to perform a gene and exon-level normalization, and signaling summarization by evaluating QC values for the array. The second software allows evaluating gene and exon expiration between samples. Parameters used for the analysis were: One-Way Between-Subject ANOVA (Unpaired); Fold Change (linear) < -2 or Fold Change (linear) > 2; and ANOVA p-value (Condition pair) < 0.05.
Fold Change (linear) < -2 or Fold Change (linear) > 2; and ANOVA p-value (Condition pair) < 0.05.
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| |
|
| Submission date |
Aug 21, 2015 |
| Last update date |
Aug 25, 2015 |
| Contact name |
Ruth Ruiz-Esparza |
| E-mail(s) |
bioruthy@yahoo.com
|
| Phone |
(55) 56590883
|
| Organization name |
Instituto Mexicano del Seguro Social
|
| Department |
Pediatría
|
| Lab |
Genética Humana
|
| Street address |
Centro Médico Nacional Siglo XXI, IMSS, México,
|
| City |
México. |
| State/province |
D.F. |
| ZIP/Postal code |
06720 |
| Country |
Mexico |
| |
|
| Platform ID |
GPL17586 |
| Series (1) |
| GSE72269 |
Differentially Expressed LncRNAs Might be Controlling Signaling Pathways by Establishing Interactions with mRNAs and MicroRNAs in Pediatric Astrocytoma |
|
| Relations |
| Alternative to |
GSM1860459 (exon-level analysis) |
