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Sample GSM102071 Query DataSets for GSM102071
Status Public on May 04, 2007
Title DFVF1_pig4_lung_infected
Sample type RNA
 
Channel 1
Source name Lung tissue, infected
Organism Sus scrofa
Characteristics Animal: 10-12 week old castrate of Danish Landrace/Yorkshire/Duroc from a high health herd (Specific Pathogen Free)
Tissue: lung
State: infected with Actinobacillus pleuropneumoniae
Biomaterial provider Danish Institute for Food and Veterinary Research
Extracted molecule total RNA
Extraction protocol RNeasy Maxi Kit with DNase treatment (Qiagen)
Label Alexa 594
Label protocol Superscript Indirect cDNA Labeling System (Invitrogen) and ARES cDNA labeling kit (Molecular Probes/Invitrogen)
 
Channel 2
Source name Common reference
Organism Sus scrofa
Characteristics Mixture of total RNA purified from infected and healthy pig lung tissue
Biomaterial provider Danish Institute for Food and Veterinary Research
Extracted molecule total RNA
Extraction protocol RNeasy Maxi Kit with DNase treatment (Qiagen)
Label Alexa 488
Label protocol Superscript Indirect cDNA Labeling System (Invitrogen) and ARES cDNA labeling kit (Molecular Probes/Invitrogen)
 
 
Hybridization protocol The slides were pre-hybridized (5 x SSC, 0.1 % SDS, 1 mg/mL BSA, 0.22 µm filtrated) at 42 ºC for 45 min and washed (2x5 dips in water) followed by a dip in isopropanol and centrifugation (300 x g). mSeries LifterSlip (25x60 mm, Erie Scientific Company) were cleaned by dipping in water followed by several dips in ethanol and left for drying. A lifterslip was placed on each slide and incubated at 42 ºC to preheat. The hybridization chamber, a 6x17x22 cm lidded plastic box with a stainless steel “table” and a napkin wetted with hybridization buffer, was placed at 42 ºC to preheat. The purified labeled cDNA were combined pair wise and 20 µg Poly-dA (Invitrogen), 25 µg yeast tRNA (Invitrogen) and 20 µg pig CoT-DNA was added. The samples were speedvaced until dry (medium heat, app. 30 min), resuspended in 70 µL hybridization buffer (50 % formamide, 5x SSC, 0.1 % SDS), incubated at 95 ºC for 3 min, snap cooled on ice and centrifuged. The samples were applied to the pre-heated slides and placed in the hybridization chamber which was sealed and incubated at 42ºC for 12 to 16 hours. Following hybridization, each array was immersed in 100 mL wash-1 (2x SSC, 0.1% SDS, 0.22 µm filtrated) at 42ºC until the lifterslip moves freely away from the slide. The arrays were transferred to a mBox (Erie Scientific Company) and immersed in wash-1 (2x SSC, 0.1% SDS, 0.22 µm filtrated) at 42ºC for 5 min., in wash-2 (0.1x SSC, 0.1% SDS, 0.22 µm filtrated) at ambient temperature for 10 min., 4 times in wash-3 (0.1x SSC, 0.22 µm filtrated) at ambient temperature for 1 min., rinsed in wash-4 (0.01x SSC, 0.22 µm filtrated) for 5 to 10 sec., dipped 5 times in water and dried by centrifugation (300 x g).
Scan protocol Scanner: ScanArray Express HT system (Perkin Elmer), 10 µm resolution, 100 % LP and PMT adjusted individually for each channel. Image analysis software: ScanArray Express, ver. 3.0 (Perkin Elmer) using the histogram method with default settings.
Description RNA from lung tissue was extracted from healthy pig lung tissue and from lung tissue infected with Actinobacillus pleuropneumoniae, reverse transcribed and labeled with Alexa 594. A common reference (equal amounts of RNA from all samples) was reverse transcribed and labeled with Alexa 488. Samples and reference were hybridized simultaneously to DIAS_PIG_27K2_v2 slides and fluorescence in the Alexa 594 and Alexa 488 channels was acquired using a microarray scanner.
Data processing Statistical analysis was carried out in the R computing environment (ver 2.3.0, developmental version, for Windows) using the package Linear Models for Microarray Analysis (Limma, ver 2.4.11) which is part of the Bioconductor project. The log2-transformed ratios of Alexa-594 to Alexa-488 (not background corrected) were normalized within-slide using printtip-loess with default parameters as implemented in Limma and can be found in the “Value” column.
 
Submission date Mar 29, 2006
Last update date May 03, 2007
Contact name Jakob Hedegaard
E-mail(s) Jakob.Hedegaard@ki.au.dk
Phone (+45)89991363
Organization name Aarhus University, Faculty of Agricultural Sciences
Department Department of Genetics and Biotechnology
Lab Molecular Genetics and System Biology
Street address PO-box 50
City Tjele
ZIP/Postal code DK-8830
Country Denmark
 
Platform ID GPL3594
Series (1)
GSE4577 Early pig response to experimental infection with Actinobacillus pleuropneomuniae

Data table header descriptions
ID_REF
VALUE Printtiploess-normalized log2 ratios (594/488)
Ch1 Mean Ch1 (Alexa 594) mean feature intensity
Ch2 Mean Ch2 (Alexa 488) mean feature intensity

Data table
ID_REF VALUE Ch1 Mean Ch2 Mean
100108.4 -0.029803 1072 1216
100108.3 0.052636 1116 1206
100120.4 -0.001184 1108 1261
100120.3 -0.174012 1022 1283
100156.4 -0.006108 1098 1241
100156.3 0.102827 1134 1182
100168.4 -0.058952 1052 1217
100168.3 -0.160942 1003 1246
100204.4 -0.228069 919 1165
100204.3 0.010224 1024 1091
100216.4 0.262578 1163 1077
100216.3 -0.107667 985 1139
100252.4 -0.007367 1076 1201
100252.3 0.087876 1073 1113
100264.4 -0.621415 1106 2757
100264.3 -0.479717 1127 2505
100300.4 0.219541 1129 1076
100300.3 0.047649 1046 1093
100312.4 -0.567236 1598 3880
100312.3 -0.436726 1690 3734

Total number of rows: 27648

Table truncated, full table size 761 Kbytes.




Supplementary file Size Download File type/resource
GSM102071_12755470.csv.gz 4.3 Mb (ftp)(http) CSV

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