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Sample GSM873267 Query DataSets for GSM873267
Status Public on Feb 10, 2012
Title 12.5dpc MC vs 13.5dpc CA (Dye Combination 1)
Sample type RNA
 
Channel 1
Source name Mesenchymal condensation, 12.5dpc hind limb
Organism Mus musculus
Characteristics tissue: Mesenchymal condensation hind limb
age: 12.5dpc
strain: Swiss white
Treatment protocol Dissected limb buds were embedded in Tissue-Tek OCT (Sakura Fine Technical), snap-frozen in isopentane, and stored at -80° C. 6um cryosections were prepared using a cryostat (Leica CM1850), mounted on RNAse-free SuperFrost Plus slides (Biolab Scientific), and processed immediately by being fixed in 70% ethanol, rinsed in RNAse-free water, and dehydrated in 70%, 95%, and 100% ethanol for 30 seconds each. They were then stored in 100% ethanol at -80° C. Immediately prior to microdissection, sections were air-dried and the tissue of interest was microdissected using an ophthalmic scalpel (Feather) fixed to the scanning xy-object guide. Light microscopy was performed using an Eclipse 80i light microscope (Nikon).
Growth protocol Pregnant Swiss white mice were sacrificed in accordance with institutional ethical guidelines at the appropriate stage of gestation. Embryos were then harvested and culled, and hind limb buds were dissected.
Extracted molecule total RNA
Extraction protocol Microdissected tissues were collected in TRIzol reagent (Invitrogen) for RNA extraction and purification, which was performed following the manufacturer’s specifications. All total RNA samples were subjected to two rounds of linear amplification using the MessageAmp II aRNA Amplification Kit (Ambion), following the manufacturer’s protocol. 100-150ng of first-round amplified aRNA was used as template for each second round amplification. Following total RNA extraction and amplification, the yield, purity, and integrity of all RNA samples were validated by capillary electrophoresis with a Bioanalyzer 2100 (Agilent Technologies), using a Series II RNA 6000 Pico Kit (Agilent Technologies), according to the manufacturer’s specifications.
Label Cy3
Label protocol Amplified RNA samples (2ug) were labelled with Cy3 or Cy5 (Amersham), according to the manufacturer’s specifications. Fluorophore incorporation and yield were determined by spectrophotometry using a Nanodrop ND-1000 spectrophotometer (Thermo Fisher Scientific).
 
Channel 2
Source name Cartilage anlagen, 13.5dpc hind limb
Organism Mus musculus
Characteristics strain: Swiss white
tissue: Cartilage anlagen hind limb
age: 13.5dpc
Treatment protocol Dissected limb buds were embedded in Tissue-Tek OCT (Sakura Fine Technical), snap-frozen in isopentane, and stored at -80° C. 6um cryosections were prepared using a cryostat (Leica CM1850), mounted on RNAse-free SuperFrost Plus slides (Biolab Scientific), and processed immediately by being fixed in 70% ethanol, rinsed in RNAse-free water, and dehydrated in 70%, 95%, and 100% ethanol for 30 seconds each. They were then stored in 100% ethanol at -80° C. Immediately prior to microdissection, sections were air-dried and the tissue of interest was microdissected using an ophthalmic scalpel (Feather) fixed to the scanning xy-object guide. Light microscopy was performed using an Eclipse 80i light microscope (Nikon).
Growth protocol Pregnant Swiss white mice were sacrificed in accordance with institutional ethical guidelines at the appropriate stage of gestation. Embryos were then harvested and culled, and hind limb buds were dissected.
Extracted molecule total RNA
Extraction protocol Microdissected tissues were collected in TRIzol reagent (Invitrogen) for RNA extraction and purification, which was performed following the manufacturer’s specifications. All total RNA samples were subjected to two rounds of linear amplification using the MessageAmp II aRNA Amplification Kit (Ambion), following the manufacturer’s protocol. 100-150ng of first-round amplified aRNA was used as template for each second round amplification. Following total RNA extraction and amplification, the yield, purity, and integrity of all RNA samples were validated by capillary electrophoresis with a Bioanalyzer 2100 (Agilent Technologies), using a Series II RNA 6000 Pico Kit (Agilent Technologies), according to the manufacturer’s specifications.
Label Cy5
Label protocol Amplified RNA samples (2ug) were labelled with Cy3 or Cy5 (Amersham), according to the manufacturer’s specifications. Fluorophore incorporation and yield were determined by spectrophotometry using a Nanodrop ND-1000 spectrophotometer (Thermo Fisher Scientific).
 
 
Hybridization protocol All aRNA samples were hybridized to 44K mouse whole genome microarrays (Agilent Technologies), in two color hybridizations including dye swaps, and in strict accordance with the manufacturer’s specifications.
Scan protocol All microarrays were scanned at 10um resolution on an Axon 4000B scanner
Data processing The microarray features were acquired using GenePix Pro 4.1 software, and the raw data was processed using a print tip loess method of normalization with limmaGUI, which is an implemented package of R used to fit linear models to microarray data.
 
Submission date Feb 09, 2012
Last update date Feb 10, 2012
Contact name Trevor L Cameron
E-mail(s) trevor.cameron@mcri.edu.au
Organization name Murdoch Childrens Research Institute
Street address Flemington Road
City Parkville
ZIP/Postal code 3052
Country Australia
 
Platform ID GPL4134
Series (1)
GSE35669 Global comparative transcriptome analysis of cartilage formation in vivo

Data table header descriptions
ID_REF
VALUE Log ratios (635/532) not normalized

Data table
ID_REF VALUE
45220 0.873
45050 0.787
44880 -1.222
44710 -0.263
44540 0
44370 0.263
44200 -0.263
44030 -1.807
43860 -0.263
43690 -2.807
43520 -1
43350 0
43180 0
43010 0
42840 -0.807
42670 -0.737
42500 -0.485
42330 -0.585
42160 0
41990 0

Total number of rows: 45220

Table truncated, full table size 519 Kbytes.




Supplementary file Size Download File type/resource
GSM873267_Cy3_12.5dpc_vs_Cy5_13.5dpc.gpr.gz 6.0 Mb (ftp)(http) GPR
Raw data provided as supplementary file
Raw data included within Sample table
Processed data not provided for this record

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