|
| Status |
Public on May 20, 2008 |
| Title |
Human Saliva Male 4 |
| Sample type |
RNA |
| |
|
| Source name |
Healthy male
|
| Organism |
Homo sapiens |
| Characteristics |
saliva supernatant
male
|
| Growth protocol |
Whole saliva was collected by direct spitting into 50ml Falcon tubes placed on ice. Supernatant was obtained following centrifugation of whole saliva for 15 minutes at 2600 g.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated from 560 µL of saliva supernatant with the RNeasy Mini Kit (Qiagen), according to the manufacturer’s instructions. Inclusion of 10 mL/L of the RNase inhibitor NucleoGuard (AmpTec) in the lysis buffer improved RNA yield and recoveries of long transcripts. All samples were treated with TURBO DNA-free (Ambion) to remove trace amounts of genomic DNA. A 2-round amplification was performed with the ExpressArt TRinucleotide mRNA Amplification Kit (AmpTec) according to the manufacturer’s instructions.
|
| Label |
biotin
|
| Label protocol |
Single-stranded cDNA was generated from the amplified cRNA with the WT cDNA Synthesis Kit (Affymetrix) and then fragmented and labeled with the WT Terminal Labeling Kit (Affymetrix).
|
| |
|
| Hybridization protocol |
Samples were hybridized with GeneChip Human Exon 1.0 ST Arrays (Affymetrix) and scanned at the UCLA Microarray Core Facility.
|
| Scan protocol |
Array scanning was performed according to the manufacturer's instruction (Affymetrix)
|
| Description |
Library files included in primary analysis:
HuEx-1_0-st-v2.r2.pgf
HuEx-1_0-st-v2.r2.clf
HuEx-1_0-st-v2.r2.antigenomic.bgp
HuEx-1_0-st-v2.r2.dt1.hg18.core.mps
|
| Data processing |
Raw data were processed with the Exon Array Computational Tool (ExACT) (Affymetrix) for background correction and normalization. Data analysis and statistical evaluations were performed with customized R codes (version 2.3.1, http://www.r-project.org/). We defined a probeset as present when it had a P value <0.001 and an Intensity value >200. These criteria were suggested by the results of preliminary experiments. The SECT includes all probesets present in at least 16 of the 18 arrays. In addition, we refined the SECT to remove GC-rich (i.e., >=80%) probesets. The concordance between the SECT and the normal salivary core transcriptome (NSCT) was evaluated assuming hypergeometric distributions.
|
| |
|
| Submission date |
May 21, 2007 |
| Last update date |
Sep 18, 2008 |
| Contact name |
David T.W. Wong |
| E-mail(s) |
dtww@ucla.edu, mikehu@ucla.edu
|
| Organization name |
University of California, Los Angeles
|
| Department |
Dental Research Institute
|
| Street address |
10833 Le Conte Ave, CHS 73-017
|
| City |
Los Angeles |
| State/province |
CA |
| ZIP/Postal code |
90095 |
| Country |
USA |
| |
|
| Platform ID |
GPL5175 |
| Series (1) |
| GSE7760 |
Exon Level Expression Profiling: a Novel Unbiased Transcriptome Analysis for Body Fluids |
|
