|
| Status |
Public on Dec 01, 2012 |
| Title |
A. pleuropneumoniae__serotype6_24h__rep1 [46-1] |
| Sample type |
RNA |
| |
|
| Source name |
Actinobacillus pleuropneumoniae 7712640 from pig lung 24h
|
| Organism |
Actinobacillus pleuropneumoniae |
| Characteristics |
serotype: 6
strain: 7712640
|
| Treatment protocol |
Prior to RNA extraction, 100 to 300 mg lung tissue with visual lesions was finely chopped by scalpel, transferred to 5 ml Phenol:Guanidinthiocyanat lysis buffer (provided in the Qiagen kit) in which it was divided further by a Tissue shearer for 2 min.
|
| Growth protocol |
Immediately post mortem, infected lung tissue was isolated, cut into pieces smaller than 0.5×0.5 cm and preserved in RNAlater stabilization reagent (Ambion, Cambridgeshire, United Kingdom) at -20°C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the RNeasy Lipid kit (QIAGEN, Hilden, Germany). Genomic DNA was eliminated by RNase-free DNase I treatment during the isolation procedure. After RNA extraction, the material was further treated by TURBO™ DNase, according to the protocol provided by the manufacturer (Ambion). The RNA concentration and quality were measured by NanoDrop (Thermo Scientific, Wilmington, DE, USA) and Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA), respectively. For each sample, 30 µg of total RNA was enriched for bacterial RNA applying the MicrobEnrich Kit according to the supplied protocol (Ambion). Subsequently, one µg of the enriched RNA was amplified using a MessageAmp II-Bacteria kit (Ambion) according to the manufacturer's instructions.
|
| Label |
Cy3
|
| Label protocol |
Labeling was performed by NimbleGen Systems Inc. (Madison, WI USA) following their standard operating protocol. See www.nimblegen.com.
|
| |
|
| Hybridization protocol |
Hybridization was performed by NimbleGen Systems Inc. (Madison, WI, USA) following their standard operating protocol. See www.nimblegen.com.
|
| Scan protocol |
Scanning was performed by NimbleGen Systems Inc. (Madison, WI USA*) following their standard operating protocol. See www.nimblegen.com.
|
| Description |
This sample is of A. pleuropneumoniae strain 7712640. It is the first of three biological replicates for pig no. 46, each isolated from separate lung lesions.
|
| Data processing |
The Robust Multichip Average (RMA)function was applied for normalization of the microarray data (Irizarry et al. Biostatistics 4(2):249). For RMA normalization we have used RGui version 2.9.2 (2009-08-24) (http://cran.r-project.org/bin/windows/base/), using the package Oligo.
|
| |
|
| Submission date |
Nov 29, 2011 |
| Last update date |
Dec 01, 2012 |
| Contact name |
Kirstine Klitgaard |
| E-mail(s) |
kksc@vet.dtu.dk
|
| Phone |
0045 35886265
|
| Organization name |
National Veterinary Institute, Technical University of Denmark
|
| Street address |
Bulowsvej 27
|
| City |
Copenhagen |
| ZIP/Postal code |
1790 |
| Country |
Denmark |
| |
|
| Platform ID |
GPL11003 |
| Series (1) |
| GSE33999 |
Genome wide transcriptional portrait of Actinobacillus pleuropneumoniae during invasive disease reveals new strategies for survival and persistence in the host |
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