|
| Status |
Public on Mar 08, 2017 |
| Title |
AorticSMC_SRFknockout_AdEmpty_rep1 |
| Sample type |
RNA |
| |
|
| Source name |
AorticSMC_SRFKO+AdEmpty
|
| Organism |
Mus musculus |
| Characteristics |
tissue type: Cultured mouse aortic smooth muscle cell
passage: P3
|
| Treatment protocol |
Adenovirus transduction with MOI=100 for 3 days before harvest.
|
| Growth protocol |
DMEM/F12 with 20%FBS for maintaining, DMEM/F12 with 10% for experiment
|
| Extracted molecule |
total RNA |
| Extraction protocol |
miR-easy kit extracted RNA (0.5 ug) was submitted for microarray.
|
| Label |
Cy3
|
| Label protocol |
Sample labeling was performed according to the Agilent One-Color Microarray-Based Gene Expression Analysis protocol (Agilent Technology) with minor modifications. Briefly, each sample was transcribed into fluorescent cRNA along the entire length of the transcripts without 3’ bias utilizing a random priming method (Arraystar Flash RNA Labeling Kit, Arraystar). The labeled cRNAs were purified by RNeasy Mini Kit (Qiagen). The concentration and specific activity of the labeled cRNAs (pmol Cy3/ug cRNA) were measured by NanoDrop ND-1000. 1 ug of each labeled cRNA was fragmented by adding 5 ul 10 × Blocking Agent and 1 ul of 25 × Fragmentation Buffer, then heated the mixture at 60 °C for 30 min, finally 25 ul 2 × GE Hybridization buffer was added to dilute the labeled cRNA.
|
| |
|
| Hybridization protocol |
50 ul of hybridization solution containing the labeled cRNA was dispensed into the gasket slide and assembled to the LncRNA expression microarray slide. The slides were incubated for 17 hours at 65°C in an Agilent Hybridization Oven. The hybridized arrays were washed and fixed.
|
| Scan protocol |
Agilent DNA Microarray Scanner (part number G2505C) was used to scan the arrays.
|
| Data processing |
Agilent Feature Extraction software (version 11.0.1.1) was used to analyze acquired array images. Quantile normalization and subsequent data processing were performed with using the GeneSpring GX v11.5.1 software package (Agilent Technologies). After quantile normalization of the raw data, LncRNAs and mRNAs that at least 4 out of 8 samples have flags in Present or Marginal (“All Targets Value”) were chosen for further data analysis.
|
| |
|
| Submission date |
Aug 23, 2016 |
| Last update date |
Mar 08, 2017 |
| Contact name |
Xiaochun Long |
| E-mail(s) |
Xlong@augusta.edu
|
| Phone |
5854290865
|
| Organization name |
Augusta University
|
| Department |
Vascular Biology Center
|
| Street address |
1460 Laney Walker Blvd, CL-3004
|
| City |
Augusta |
| State/province |
GA |
| ZIP/Postal code |
30912 |
| Country |
USA |
| |
|
| Platform ID |
GPL19286 |
| Series (1) |
| GSE85930 |
LncRNA and mRNA Expression in aortic smooth muscle cells with Srf knockout or Myocd over-expression |
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