Dataset Information

Investigating the effect of CodY mutation on the gene expression in Streptococcus suis serotype 2 SC19 strain.

ABSTRACT: To investigate the effect of CodY mutation on the gene expression in Streptococcus suis serotype 2 SC19 strain, we have employed whole genome microarray expression profiling as a discovery platform to identify genes regulated by CodY mutation. DNA microarray analysis was performed using an Agilent custom-designed oligonucleotide microarray. Based upon the whole genome sequence of SC19 , specific 60-mer oligonucleotide probes were designed using eArray (https://earray.chem.agilent.com/earray/), to cover all annotated genes. Probes were printed seven times on microarray slides. Three biological replicates of total RNA from two wild type strains and from two codY mutant strains were amplified and labeled with Cy3-CTP using Low Input Quick Amp Labeling Kit, one-color(Agilent technologies, US), following the manufacturer’s instructions. Labeled cRNA was purified using the RNeasy mini kit (Qiagen). After fragmentation, microarray slides were hybridized with 600 ng Cy3-labeled cRNA. Hybridization was performed at 65 °C for 17 h with rotation at 10 rpm. Microarray slides were washed and scanned by an Agilent Microarray Scanner (G2565BA). Those genes with greater than two-fold change ratios were regarded as differentially expressed genes.

ORGANISM(S): Streptococcus suis

PROVIDER: GSE68068 | GEO | 2015/04/21

SECONDARY ACCESSION(S): PRJNA281651

REPOSITORIES: GEO

ACCESS DATA

Similar Datasets

Project description:Surgically resected gastric adenocarcinoma samples and their paired disease-free (R0) marginal nonmalignant tissue were obtained from 14 patients with gastric adenocarcinoma. RNA was isolated using the RNAeasy Kit (Qiagen Valencia, CA) from 15 mg of tissue. The quality of total RNA and its integrity was assessed using the Bioanalyzer 2100 (Agilent, Palo Alto, CA) and RIN value (RNA Integrity Number) was recorded for all the samples for intact 18S and 28S rRNA. Total RNA (800 ng) from each sample was reverse transcribed and linear amplified using the low RNA input linear amplification kit (Agilent Technologies). After synthesis of the first and second strand of cDNA, the product was used in an in vitro transcription reaction to generate cRNAs in the presence of cyanine 3-labeled UTP (Cy3) for normal and cyanine 5-labeled UTP (Cy5) for tumor. Fragmented Cy3-labeled cRNA of the control sample were mixed with equal amounts of Cy5-labeled cRNA from the gastric tumor sample, and the mixtures were hybridized onto 44K whole human genome DNA microarrays (Agilent Technologies) for 17 hrs at 65°C with constant rotation (10 rpm). Subsequently, the arrays were washed according to manufacturer’s instructions. The slides were scanned using an Agilent microarray scanner (G2565AA), and the images were processed using the Agilent feature extraction software AFE 9.5. GeneSpring GX v10.1 (Agilent Technologies) was used to analyze the expression profiles obtained after microarray hybridization. The feature extraction files from all the samples were uploaded to GeneSpring. Following intensity-dependent normalization (Lowess), the data was subjected to statistical analysis. A two tailed student’s t-test was used to determine significance of the differences observed between the normal and tumor samples. Benjamini Hochberg algorithm was used to compute false discovery rates. Genes were filtered based on P value threshold of 0.001 and false discovery rate of less than 1%.

Dataset Information

Investigating the effect of CodY mutation on the gene expression in Streptococcus suis serotype 2 SC19 strain.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure