|
| Status |
Public on Oct 02, 2006 |
| Title |
KO - knock-out Treated - 12 hrs - repeat 2 - Cy5 - mAdbID:61760 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
WT Primary Hepatocytes - Pooled -Cy3
|
| Organism |
Mus musculus |
| Characteristics |
Strain: C57BL
Gender: male
Age: 9-10 weeks
Tissue: liver
Developmental stage: adult
Cell type: Primary hepatocytes
|
| Growth protocol |
Cell Culture Protocol
Media: 1:1 DMEM and HAM's F12 supplemented with ITS
Other: Primary hepatocytes were seeded at 2x106 in 10 cm dishes in the plating medium supplemented with 10% FBS . After 4 hours, the plating medium was replaced to serum-free medium. The following day cells were treated with 50 ng/ml of rhHGF for 0.5, 2, 12 and 24 h. Triplicate cell cultures were established from three individual mice.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Extraction Protocol
Extraction method: Trizol protocol
|
| Label |
cy3
|
| Label protocol |
Cy3 Sample Labeling Protocol - generic
|
| |
|
| Channel 2 |
| Source name |
KO - Met knock-out -treated 12 hr - Cy5
|
| Organism |
Mus musculus |
| Characteristics |
Strain: 129SV/C57BL6
Gender: male
Age: 9-10 weeks
Tissue: liver
Developmental stage: adult
Cell type: Primary hepatocytes
Genotype/Genetic variation gene knock out
Genotype/Genetic variation: Alb-Cre +/-; Met Fl/Fl
|
| Treatment protocol |
Treatment type: compound
Agent: HGF (Hepatocyte Growth Factor)
Treatment dose: 50 nM
Treatment time: 0.5 hour
|
| Growth protocol |
Cell Culture Protocol
Media: 1:1 DMEM and HAM's F12 supplemented with ITS
Other: Primary hepatocytes were seeded at 2x106 in 10 cm dishes in the plating medium supplemented with 10% FBS . After 4 hours, the plating medium was replaced to serum-free medium. The following day cells were treated with 50 ng/ml of rhHGF for 0.5, 2, 12 and 24 h. Triplicate cell cultures were established from three individual mice.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Extraction Protocol
Extraction method: Trizol protocol
|
| Label |
cy5
|
| Label protocol |
Cy5 Sample Labeling Protocol - generic
|
| |
|
| |
| Hybridization protocol |
NCI Oligo Microarray Hybridization
Other: For pre-hybrization, apply 40 ul of pre-hybridization buffer (5X SSC, 0.1% SDS, 1% BSA) to the array and incubate at 42øC for at least 30 minutes and up to an hour. Wash off the pre-hybridization solution by rapidly plunging the slide in distilled water for 2 minutes, then transfer slide to 100% isopropanol for 2 minutes. Allow slide to air dry completely prior to use. (Can spin dry if in a rush.) (NOTE: Do not exceed 1 hour after pre-hybridization/drying before setting up hybridization.) For hybridization, combine Cy3 and Cy5 labeled targets together (~9 ul recovered for each). Add 1 ul COT-1 DNA (8-10 ug/ul) and 1 ul poly A (8-10 ug/ul). Denature target at 100øC for 1 minute, then snap cool on ice. (Final volume should be about 20 ul.) Make fresh 2X Formamide hybridization buffer (50% formamide, 10X SSC, 0.2% SDS) and warm to 42øC just before adding to samples. Add 20 ul of 2X F-hyb buffer to samples. Load 40 ul sample onto microarray. Add 20 ul of 3X SSC to wells in hyb chamber to maintain humidity. Incubate overnight (12-16 hours) at 42øC in water bath or hybridization oven. After hybridization of slides, wash slides for 2 minute in 2X SSC with 0.1% SDS (with occasional plunging), for 2 minute in 1X SSC (with occasional plunging), for 2 minutes in 0.2X SSC (with occasional plunging), and spin for 3 minutes at 650 rpm to dry. (Refer to "NCI Microarray Manual")
|
| Description |
mAdb experiment ID: 61760
|
| Data processing |
After background correction and removal of flagged values, log base 2 expression ratios were median centered and linear transformed to obtain the log and linear values given in the data table.
|
| |
|
| Submission date |
Mar 13, 2006 |
| Last update date |
Oct 02, 2006 |
| Contact name |
Snorri S Thorgeirsson |
| E-mail(s) |
[email protected]
|
| Phone |
(301) 496-5688 x204
|
| Fax |
(301) 496-0734
|
| URL |
http://neoplasia.nci.nih.gov/
|
| Organization name |
National Cancer institute/NIH
|
| Department |
Laboratory of Experimental Carcinogenesis
|
| Lab |
Cellular and Molecular Biology Section
|
| Street address |
37 Convent Dr. # 4146
|
| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
| |
|
| Platform ID |
GPL1536 |
| Series (1) |
| GSE4451 |
Met Expression Signature in Hepatocytes |
|
| Data table header descriptions |
| ID_REF |
NCI mAdb well id plus replicate number |
| VALUE |
same as UNF_VALUE but with flagged values removed |
| PRE_VALUE |
Calibrated Ratio (CY5 channel/CY3 channel) |
| Slide_block |
Array block location |
| Slide_column |
Array column location |
| Slide_row |
Array row location |
| CY5_mean |
Red Channel Sample mean Signal (Background Subtracted) |
| CY5_SD |
Red Channel Sample Standard Deviation |
| CY5_BKD_median |
Red Channel Sample median Background Level |
| CY5_BKD_SD |
Red Channel Sample Background Standard Deviation |
| CY3_mean |
Green Channel Sample mean Signal (Background Subtracted) |
| CY3_SD |
Green Channel Sample Standard Deviation |
| CY3_BKD_median |
Green Channel Sample median Background Level |
| CY3_BKD_SD |
Green Channel Sample Background Standard Deviation |
| Flag |
Quality flag 0->good, -50->Not found, -100->Bad |
| UNF_VALUE |
log ratio (log2 of PRE_VALUE) |
