|
| Status |
Public on May 16, 2014 |
| Title |
AS-1-3 |
| Sample type |
RNA |
| |
|
| Source name |
WI38 cells / MALAT1 antisense oligonucleotide (AS-1)
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: WI38 cells (normal human diploid fibroblasts)
growth media: DMEM containing high glucose + 10% FBS, and 1% non-essential amino acid (NEA)
transfected with: MALAT1 antisense oligonucleotide (AS-1)
|
| Treatment protocol |
WI-38 cells were reverse transfected in triplicate with either control (scrambled antisense oligo) or two independent MALAT1 antisense oligonucleotides(AS-1, AS-2) at a final concentration of 100 nM using Lipofectamine RNAiMax (Invitrogen, USA).
|
| Growth protocol |
WI-38 cells were grown in DMEM containing high glucose + 10% FBS, and 1% non-essential amino acid (NEA).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using RNeasy kit (Qiagen) according to the manufacturer’s instructions. Quality and quantity of the total RNA was checked with the Agilent 2100 bioanalyzer using RNA 6000 Nano chips.
|
| Label |
Streptavidin-Cy3 bound to biotin
|
| Label protocol |
Standard Illumina protocol using Illumina TotalPrep RNA Amplification Kit (Ambion; Austin, TX, cat # IL1791) In short, 0.5g of total RNA was first converted into single-stranded cDNA with reverse transcriptase using an oligo-dT primer containing the T7 RNA polymerase promoter site and then copied to produce double-stranded cDNA molecules. The double stranded cDNA was cleaned and concentrated with the supplied columns and used in an overnight in-vitro transcription reaction where single-stranded RNA (cRNA) was generated and labeled by incorporation of biotin-16-UTP.
|
| |
|
| Hybridization protocol |
Standard Illumina protocol. In short, a total of 0.75ug of biotin-labeled cRNA was hybridized at 58 degrees C for 16 hours to Illumina's Sentrix Human HT-12 v4 Expression BeadChips (Illumina, San Diego, CA). Each BeadChip has ~47,000 annotated RefSeq transcripts with approximately 15-fold redundancy. The arrays were washed, blocked and the biotin labeled cRNA was detected by staining with streptavidin-Cy3.
|
| Scan protocol |
Arrays were scanned at a resolution of 0.8um using the Beadstation 500 X from Illumina.
|
| Description |
WI-38 cells reverse transfected with MALAT1 antisense oligonucleotide,(AS-1) final concentration 100 nM, Replicate #3
|
| Data processing |
Data was extracted using the Illumina GenomeStudio software(v1.6.0). Any spots at or below the background were filtered out using an Illumina detection p value of 0.02 and above. The natural log of all remaining scores were used to find the avg and std of each array and the z-score normalization was calculated and presented below. Z-score = (raw value - avg)/std. Complete data including detection scores will be included in the supplemental file.
|
| |
|
| Submission date |
Feb 11, 2013 |
| Last update date |
Jun 22, 2020 |
| Contact name |
Supriyo De |
| Organization name |
NIA-IRP, NIH
|
| Department |
Laboratory of Genetics and Genomics
|
| Lab |
Computational Biology & Genomics Core
|
| Street address |
251 Bayview Blvd
|
| City |
Baltimore |
| State/province |
Maryland |
| ZIP/Postal code |
21224 |
| Country |
USA |
| |
|
| Platform ID |
GPL10558 |
| Series (1) |
| GSE44240 |
Long Noncoding RNA MALAT1 Controls Cell Cycle Progression by Regulating the Expression of Oncogenic Transcription Factor B-MYB |
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