Project description:Array Design; Mouse Compugen 22K Spotted Oligo Author; Sean Grimmond Common Reference (Yes/No, ID); Yes Mouse universal Reference (Stratagene) Concentration; 100 Data processing/normalization; Lowess Normalisation GeneSpring) Developmental Stage; embryonic Diseased/Normal; normal Dye Swap; red/green Experiment Type; time course Genetic Characteristic; Wild type Image Analysis Software; Imagene5.5 (Biodiscovery) Labeling Protocol; Amino Allyl Message Amp- Ambion Organ/Organism part; kidney Organism; Mus Musculus RNA type; aRNA Message Amp-ambion Tissue Type; Pool of whole organs Organization; IMB, Uni of QLD, Brisbane Australia Sample Source; primary tissue Sampling Method; Dissection of whole organ Sex; Pool of both sexes Strain/Cell-line; CD1 Series_overall_design: To account for dye swap, the signal channel and control channel measurements for each sample were reversed. A Lowess curve was fit to the log-intensity versus log-ratio plot. 20.0% of the data was used to calculate the Lowess fit at each point. This curve was used to adjust the control value for each measurement. If the control channel was lower than 10 then 10 was used instead. Specific samples were normalized to one another: sample(s) 1-38 were normalized against the median of the control sample(s) 1-38. Each measurement for each gene in those specific samples was divided by the median of that gene's measurements in the corresponding control samples. Keywords: time-course
Project description:Purpose: The goal of this study is to identify vasopressin-regulated genes in mouse kidney collecting duct cell line mpkCCD. To explore dynamic regulation of vasoressin-mediated transcriptional regulation, transcriptome of vasopressin-treated cells at four different time points (3h, 6h, 12h, and 24h) were profiled and analyzed. Methods: Total RNAs were isolated from vasopressin-treated mpkCCD cells at different time points (3h, 6h, 12h, and 24h). Three replicates for vehicle- or vasopressin-treated group were generated at each tested time point. cDNA libraries were prepared using a Nextera DNA library preparation protocol. The sequence reads from Illumina HiSeq3000 platform were qualified and quantified at the transcript level using Salmon (0.14.1). Differential expression analysis were performed using edgeR. Results: mRNA profiles of mouse kidney collecting duct mpkCCD cells treated with vasopressin analog (dDAVP) for four different time potins (3h, 6h, 12h, and 24h) were generated using an optimized RNA-Seq workflow. Transcript level quantification using a pseudo-alignment quantification method (Salmon) was performed to calculate transcript abundance in each sample. Then differential expression analysis identified the differentially expressed genes including Aqp2 gene at each time point comparison (dDAVP vs vehicle, FDR < 0.05). In addition, several new vasopressin-responsive genes that have not been elucidated before were identified. Conclusions: This study revealed dynamic changes of vasopressin-responsive gene expression within 24 hours in mouse kidney collecting duct cells. The results from time-course transcriptome profiling identified the known vasopressin-responsive genes and several novel gene that are regulated temporally.
Project description:Array Design; Mouse Compugen 22K Spotted Oligo Author; Sean Grimmond Common Reference (Yes/No, ID); Yes Mouse universal Reference (Stratagene) Concentration; 100 Data processing/normalization; Lowess Normalisation GeneSpring) Developmental Stage; embryonic Diseased/Normal; normal Dye Swap; red/green Experiment Type; time course Genetic Characteristic; Wild type Image Analysis Software; Imagene5.5 (Biodiscovery) Labeling Protocol; Amino Allyl Message Amp- Ambion Organ/Organism part; kidney Organism; Mus Musculus RNA type; aRNA Message Amp-ambion Tissue Type; Pool of whole organs Organization; IMB, Uni of QLD, Brisbane Australia Sample Source; primary tissue Sampling Method; Dissection of whole organ Sex; Pool of both sexes Strain/Cell-line; CD1 Series_overall_design: To account for dye swap, the signal channel and control channel measurements for each sample were reversed. A Lowess curve was fit to the log-intensity versus log-ratio plot. 20.0% of the data was used to calculate the Lowess fit at each point. This curve was used to adjust the control value for each measurement. If the control channel was lower than 10 then 10 was used instead. Specific samples were normalized to one another: sample(s) 1-38 were normalized against the median of the control sample(s) 1-38. Each measurement for each gene in those specific samples was divided by the median of that gene's measurements in the corresponding control samples. Keywords: time-course
Project description:8 week-old male C57BL6J mice were given Gram-negative endotoxin (LPS O111:B4, 10 mg/kg) intraperitoneally at time 0. 18 hrs thereafter, they were administered 10 ml/kg 0.9% saline. Mice were sacrificed at 0, 18, or 42 hrs after LPS challenge. Kidneys were immediately collected into TRIzol for RNA preparation. Renal function was measured on blood collected at the time of tissue harvest At t=0hr, mice had normal baseline renal function. At t=18hr, mice exhibited early renal injury, At t=42hr, mice had either recovered normal renal function or had persistent renal injury. We collected kidneys from 3 mice per time point. For the 42 hr time point, we collected kidneys from 3 mice with recovered renal function and kidneys from 3 mice with persistent renal injury.
Project description:8 week-old male C57BL6J mice were given Gram-negative endotoxin (LPS O111:B4, 10 mg/kg) intraperitoneally at time 0. 18 hrs thereafter, they were administered 10 ml/kg 0.9% saline. Mice were sacrificed at 0, 18, or 42 hrs after LPS challenge. Kidneys were immediately collected into TRIzol for RNA preparation. Renal function was measured on blood collected at the time of tissue harvest At t=0hr, mice had normal baseline renal function. At t=18hr, mice exhibited early renal injury, At t=42hr, mice had either recovered normal renal function or had persistent renal injury. We collected kidneys from 3 mice per time point. For the 42 hr time point, we collected kidneys from 3 mice with recovered renal function and kidneys from 3 mice with persistent renal injury. Mouse kidneys selected at successive stages of renal injury and recovery following systemic LPS challenge and volume resuscitation following LPS challenge.