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| Status |
Public on May 30, 2014 |
| Title |
IDGSW3_vehicle_rep2_d35 |
| Sample type |
SRA |
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| Source name |
IDGSW3_vehicle_d35
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| Organism |
Mus musculus |
| Characteristics |
cell line: IDG-SW3
cell type: osteocytic cells
differentiation day: d35
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| Treatment protocol |
Basal mRNA samples at days 3, 7, 14, 21, 28, and 35 were not treated with any compound or vehicle. The vitamin D experiments were treated with 100nM of 1,25(OH)2D3 (or ethanol vehicle) for 24 hours prior to RNA harvesting.
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| Growth protocol |
Custom αMEM powdered tissue culture medium was purchased from Cellgro (Manassas, VA) and supplemented with 2.20g/L NaHCO3 (S233-3) (Fisher Scientific, Waltham, MA). Fetal Bovine Serum (FBS) (SH30396) was purchased from HyClone (Logan, Utah) and heat inactivated by incubation at 55°C for 30 minutes. Penicillin-Streptomycin Solution (SV30010) was purchased from Fisher Scientific, (Waltham, MA) and recombinant mouse interferon-gamma (rmIFN-γ) from Gibco/Invitrogen (Camarillo, CA). 10cm and 6 well collagen IV coated plates were purchased from BD Biosciences (San Jose, CA). IDG-SW3 cells (6) were expanded in 10cm collagen coated at 33°C in αMEM with 10% heat inactivated FBS, 100 units/mL penicillin/streptomycin, and 50U/mL IFNγ. Cells were harvested with 0.05% Trypsin/0.53mM EDTA (Fisher Scientific, Waltham, MA) and plated in 6 well collagen coated plates at 40,000 cells/cm2. To induce osteocytogenesis, two days after plating, cells were transferred to 37°C and media was changed to osteogenic media (αMEM with 10% heat inactivated FBS, 100 units/mL penicillin/streptomycin, 50μg/mL ascorbic acid (A4034, Sigma, St. Louis, MO) and 4mM β-glycerol phosphate (G9422, Sigma, St. Louis, MO) without IFNγ. Osteogenic media was changed three times weekly.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Directional RNA-seq libraries were prepared with the Epicentre (Madison, WI) ScriptSeq v2 RNA-seq Library Preparation Kit per manufacturer’s instructions. Ribosomal RNA depletion (RNA 6000 Nano Kit) and final library quality (High Sensitivity DNA kit) was analyzed with the Agilent Bioanalyzer (Agilent, Santa Clara, CA).
RNA was isolated using the TRI-Regent protocol (MRC, Cincinnati, OH) with an additional LiCl extraction. Eight μg of total RNA was DNase1 treated (Invitrogen) then ribosomal RNA was depleted using the RiboMinus Eukaryote Kit (Invitrogen, Carlsbad, CA) according to manufacturer’s instructions
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
Sample 26
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| Data processing |
TopHat with default settings and sample type "fr-secondstrand" for directional libraries
ArrayStar (DNASTAR) with default RNA sequencing settings. Imported BAM files from TopHat output.
Genome_build: mm9
Supplementary_files_format_and_content: tab deliminated text files include RPKM values for each sample
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| Submission date |
Feb 07, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Mark B Meyer |
| E-mail(s) |
markmeyer@wisc.edu
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| Phone |
608-890-0857
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| Organization name |
University of Wisconsin-Madison
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| Department |
Nutritional Sciences
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| Lab |
Meyer Lab
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| Street address |
1415 Linden Dr.
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| City |
Madison |
| State/province |
WI |
| ZIP/Postal code |
53706 |
| Country |
USA |
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| Platform ID |
GPL13112 |
| Series (2) |
| GSE54783 |
The Osteoblast to Osteocyte Transition: Epigenetic Changes and Response to the Vitamin D3 Hormone [RNA-seq] |
| GSE54784 |
The Osteoblast to Osteocyte Transition: Epigenetic Changes and Response to the Vitamin D3 Hormone |
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| Relations |
| BioSample |
SAMN02630890 |
| SRA |
SRX467413 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1323968_IDGSW3D35_Veh_B_RPKM.txt.gz |
117.5 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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