8 protocols
AccessionNameType
P-AFFY-6
feature_extraction
Title: Affymetrix CEL analysis. Description:
P-AFFY-2
labeling
RNA was isolated using the RNeasy Plant Mini Protocol (QIAGEN, Clifton Hill, Australia) and the quality of the RNA was assessed using the Agilent Bioanalyzer (Agilent Technologies, Palo Alto, CA) where spectrophotometric analysis was carried out to determine the A260 to A280 ratio and the concentration. For qRT-PCR, cDNA synthesis was carried out using the Biorad (Perth) iScript cDNA synthesis kit according to manufacturers instructions. For arrays, the cRNA was prepared from 3 ug of total RNA almost exactly as described in the Affymetrix GeneChip Expression Analysis Technical Manual.
P-AFFY-3
hybridization
Title: GeneChip hybridization. Description:
P-MTAB-19305
normalization data transformation protocol
Exons of the Core Meta-Probeset were used. Quantile Normalisation and RMA background correction was performed. Data analysis performed with Partek Genomics Suite, version 6.3, Copyright 2007, Partek Inc., St. Charles, MO, USA.
P-MTAB-19306
sample pooling
Three arrays (triplicate) were run for each treatment group. For each array equal quantities of RNA from 6 mice was pooled.
P-MTAB-19302
nucleic acid extraction protocol
Tissue was submerged in 1.5ml AllProtect solution (Qiagen) at autopsy. Individual colonic tissue samples (20mg) were homogenized using Lysing Matrix D tubes (MP Biomedicals) with a FastPrep-24 homogenizer (MP Biomedicals) (4.0 M/sec). RNA was extracted from the supernatant using AllPrep DNA/RNA/protein column kits according to manufacturer’s instructions (Qiagen). (Parameters: Extracted product = total_RNA, Amplification = none)
P-MTAB-19304
treatment protocol
The experiment was done on two strains of mice; AKR and BALB/c mice strains (Charles River Ltd, UK). For each strain there was an infected group and naïve control group. 6-to-8 week old male AKR and BALB/c mice strains (Charles River Ltd, UK) were used in the experiment. Infected animals received x300 Trichuris muris ova, in 200uls water, by oral gavage. 35 days later mice were killed with CO2 and colonic tissue adjacent to the ileo-caecal valve collected. This tissue was submerged in 1.5ml AllProtect® solution (Qiagen) at autopsy. Individual colonic tissue samples (20mg) were homogenized using Lysing Matrix D tubes® (MP Biomedicals) with a FastPrep®-24 homogenizer (MP Biomedicals) (4.0 M/sec). RNA was extracted from the supernatant using AllPrep® DNA/RNA/protein column kits according to manufacturer’s instructions (Qiagen). Three arrays (triplicate) were run for each treatment group. For each array equal quantities of RNA from 6 mice was pooled.
P-MTAB-19303
growth protocol
All mice were housed with free access to food and water under specific pathogen free conditions at the University of Manchester. Temperature 20-22 °C, Humidity 40-60%, Specified mouse pathogen free conditions, 12 hours light/dark cycle. Nutrients Rat and Mouse Standard Diet no.1 expanded (Ban Kingman). All experiments were performed under regulations of The UK Home Office Scientific Procedures Act (1986). 6-to-8 week old male AKR and BALB/c mice strains (Charles River Ltd, UK) were used for the experiment. (Parameters: time unit = seconds, temperature unit = C)