Computational protocol: Recovery from an Acute Infection in C. elegans Requires the GATA Transcription Factor ELT-2

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Protocol publication

[…] Total RNA was assessed for quality with an Agilent 2100 Bioanalyzer G2939A (Agilent Technologies, Santa Clara, CA) and a Nanodrop 8000 spectrophotomer (Thermo, Wilmington, DE). 100 ng of total RNA was converted to 1.65 µg Cy-3-labeled, linearly amplified cRNA using the Low Input Quick Amp (LIQA) Labeling One-Color Microarray-Based Gene Expression Analysis Kit (Agilent Technologies, Santa Clara, CA). cRNA was fragmented and added to 44 K feature Agilent C. elegans Gene Expression Microarray V2 slides (Agilent Technologies, Santa Clara, CA). Hybridization was performed in the Agilent rotisserie Hybridization Oven for 17 hours. Arrays were subsequently washed and scanned with the Agilent B scanner according to standard Agilent protocols (Agilent Technologies, Santa Clara, CA). Scanned data was log2 transformed and quantile normalized using Partek Genomics Suite (St. Louis, MO). Analysis of variance (ANOVA) t tests and fold-change calculations were also performed using Partek Genomics Suite (St. Louis, MO). For each of the 5 time points, 2 biological replicates were assessed. The microarray data was deposited in the Gene Expression Omnibus database: GSE54212. [...] Gene lists were culled from the literature and passed through WormBase Converter using the WS220 genome release as the output (references are noted in ). A total of 20,834 WS220 genes are represented by 1 or more probes in the Agilent C. elegans V2 array (Agilent Technologies, Santa Clara, CA). Gene ontology analysis was performed using the DAVID Bioinformatics Database (david.abcc.ncifcrf.gov/). The most significant gene ontology term in each DAVID functional annotation cluster was set as the significance of the overall cluster. Statistical significance of the overlap between two gene sets was calculated using the following on-line program: nemates.org/MA/progs/overlap_stats.html. Representation Factor represents the number of overlapping genes divided by the expected number of overlapping genes drawn from 2 independent groups. A background gene list of 20,834 was used for the calculation. P values were calculated using the hypergeometric probability. 1.5 kb cis-regulatory sequences were identified using WormMart (wormbase.org). Expression patterns were determined using WormMine (wormbase.org). Detailed bioinformatics protocols are available upon request. […]

Pipeline specifications

Software tools Partek Genomics Suite, WormBase Converter, DAVID
Databases GEO WormBase
Applications Gene expression microarray analysis, Transcription analysis
Organisms Salmonella enterica, Caenorhabditis elegans
Diseases Bacterial Infections, Infection