|
| Status |
Public on Feb 19, 2009 |
| Title |
HCT116 Dicer-/- #2, mock vs. HCT116 Dicer-/- #2 miR-106a, 25nM, 10hr |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
HCT116 Dicer -/-
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: HCT116 Dicer-/- #2
|
| Treatment protocol |
mock
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Qiagen RNeasy spin columns with DNAse treatment
|
| Label |
Cy3
|
| Label protocol |
custom automated version of the aminoallyl MessageAmp II kit from Ambion.
|
| |
|
| Channel 2 |
| Source name |
HCT116 Dicer -/-
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: HCT116 Dicer-/- #2
|
| Treatment protocol |
miR-106a, 25nM, 10hr
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Qiagen RNeasy spin columns with DNAse treatment
|
| Label |
Cy5
|
| Label protocol |
custom automated version of the aminoallyl MessageAmp II kit from Ambion.
|
| |
|
| |
| Hybridization protocol |
Microarrays are incubated at 40°C for 48 hours in a rotating carousel. Hybridizations to custom Agilent microarrays are completed as previously described (Hughes et al.Nat Biotech (2001), 19(4):342-7). Microarrays are washed to remove non-specific hybridized sample. Afterwards, microarrays are dried in an ozone-free nitrogen chamber.
|
| Scan protocol |
Microarrays are scanned using the Agilent LP2 laser scanner. The scanner output is a Tiff file, which contains the quantitative hybridization data from each individual microarray. The Tiff files are then processed using Rosetta custom feature extraction software.
|
| Description |
microRNAs in the miR-106b family are overexpressed in multiple tumor types and are correlated with the expression of genes that regulate the cell cycle. Consistent with these observations, miR-106b family gain of function promotes cell cycle progression, whereas loss of function reverses this phenotype. Microarray profiling uncovers multiple targets of the family, including the cyclin-dependent kinase inhibitor p21/CDKN1A. We show that p21 is a direct target of miR-106b and that its silencing plays a key role in miR-106b-induced cell cycle phenotypes. We also show that miR-106b overrides a doxorubicin-induced DNA damage checkpoint. Thus, miR-106b family members contribute to tumor cell proliferation in part by regulating cell cycle progression and by modulating checkpoint functions.
|
| Data processing |
Data were processed using the Rosetta Resolver® system. Rosetta's custom feature extraction software performs error modeling before data are loaded into the Resolver system. The Resolver system performs a squeeze operation that combines replicates of the same sequence in an array while applying error weighting. The error weighting consists of adjusting for additive and multiplicative noise. A P-value is generated and propagated throughout the system. The P-value represents the probability that a gene is expressed. The Resolver system allows users to set thresholds, below which genes of a P-value are considered to be significantly expressed. The Resolver system also combines multiple arrays using a squeezing process. If multiple spots reference one sequence, summarization is performed using an error-weighted average as described in Roland Stoughton and Hongyue Dai, Statistical Combining of Cell Expression Profiles. US Patent #6,351,712, February 26, 2002.
|
| |
|
| Submission date |
Feb 13, 2009 |
| Last update date |
Jun 25, 2009 |
| Contact name |
Irena Ivanovska |
| E-mail(s) |
irena_ivanovska@merck.com
|
| Phone |
206-802-7364
|
| Organization name |
Merck
|
| Street address |
401 Terry Ave N
|
| City |
Seattle |
| State/province |
WA |
| ZIP/Postal code |
98109 |
| Country |
USA |
| |
|
| Platform ID |
GPL4372 |
| Series (1) |
| GSE14831 |
MicroRNAs in the miR-106b family regulate p21/CDKN1A and promote cell cycle progression. |
|
