|
| Status |
Public on Jul 25, 2016 |
| Title |
HeLa-S3_siLATS1/LATS2 |
| Sample type |
RNA |
| |
|
| Source name |
HeLa-S3 cell, siLATS1/LATS2
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: HeLa-S3
tissue: cervix
|
| Treatment protocol |
siRNAs for LATS1 and/or LATS2 were transfected in HeLa-S3 cells using Lipofectamine 2000 (Life technologies), then cells were cultured for 48h prior to RNA sampling.
|
| Growth protocol |
HeLa-S3 cells were cultured in DMEM + 10% fetal bovine serum (FBS), penicillin (100U/mL), and streptomycin (100μg/mL) at 37°C, 5% CO2.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from cultured cells with miRNeasy Mini Kit (Qiagen) and processed according to the manufacturer’s protocol. The quality of the RNAs was estimated by using the RNA 6000 Nano LabChip Kit (p/n 5065-4476) on the Agilent 2100 Bioanalyzer (G2940BA; Agilent Technologies, Santa Clara, CA).
|
| Label |
Cy3
|
| Label protocol |
100 ng of total RNAs were independently reverse-transcribed using oligo-dT primers containing the T7 RNA polymerase promoter sequence to generate cDNAs and AffinityScript, RTase, which were then subjected to in vitro transcription using T7 RNA polymerase to label the cRNAs with Cy3-CTP (Amersham Pharmacia Biotech, Piscataway, NJ) using a Low input Quick-Amp Labeling Kit (Agilent Technologies).
|
| |
|
| Hybridization protocol |
Before hybridization, 1650 ng labeled cRNA of each product was fragmented and mixed with control targets and hybridization buffer according to the manufacturer's protocol (Agilent Technologies). Hybridizations to Agilent-014850 arrays were done for approximately 17h at 65°C. The slides were washed according to the manufacturer's manual.
|
| Scan protocol |
Scanning of microarrays was performed with 5µm resolution using a DNA microarray laser scanner (Agilent Technologies). The array was scanned using Agilent G2505C DNA microarray scanner. Images were quantified using Agilent Feature Extraction Software (version 10.5.1.1).
|
| Description |
HeLa-S3 cell, siLATS1/LATS2
|
| Data processing |
Agilent Feature Extraction Software (v 10.5.1.1) was used for background subtraction, LOWESS normalization, and dye-normalization.
|
| |
|
| Submission date |
Nov 21, 2014 |
| Last update date |
Jul 25, 2016 |
| Contact name |
Daisuke Okuzaki |
| E-mail(s) |
dokuzaki@biken.osaka-u.ac.jp
|
| Phone |
+81-6-6879-4935
|
| Organization name |
Osaka univ.
|
| Department |
Immunology Frontier Research Center
|
| Lab |
Human Immunology （Single Cell Genomics)
|
| Street address |
Yamadaoka 3-1
|
| City |
Suita |
| State/province |
Osaka |
| ZIP/Postal code |
565-0871 |
| Country |
Japan |
| |
|
| Platform ID |
GPL10332 |
| Series (2) |
| GSE63533 |
Transcriptome profiling of HeLa-S3 cells treated with siRNAs targeting LATS1 and/or LATS2 |
| GSE63538 |
LATS2 regulates repressive epigenetic integrity via regulation of Polycomb repressive complex 2 |
|
