Transcriptional profiling of estrogen-responsive uterine genes in Rptorf/f Pgrcre/+ mice
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE312338 | GEO | 2026/05/15
REPOSITORIES: GEO
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Project description:Purpose:We found that fat-1 transgenic bovine fetal fibroblasts have changes in the expression of energy metabolism-related genes compared with wild-type, so whether this change is due to changes in DNA methylation needs to be determined. Methods:Total DNA was extracted using QIAamp Fast DNA Tissue Kit (Qiagen, Dusseldorf, Germany) following the manufacturer's procedure. The quantity of DNA was measured by reading A260/280 ratios by spectrophotometer. When A260/280 ratios located range 1.8 to 2.0, DNA was available. The fragmented DNA samples by using sonication were subjected to bisulfite conversion. The Accel-NGS Methyl-Seq DNA Library Kit (Swift, MI, USA) was utilized for attaching adapters to single-stranded DNA fragments. Briefly, the Adaptase step is a highly efficient, proprietary reaction that simultaneously performs end repair, tailing of 3 ends, and ligation of the first truncated adapter complement to 3 ends. The Extension step is used to incorporate truncated adapter 1 by a primer extension reaction. The Ligation step is used to add the second truncated adapter to the bottom strand only. The Indexing PCR step increases yield and incorporates full length adapters. Bead-based SPRI clean-ups are used to remove both oligonucleotides and small fragments, as well as to change enzymatic buffer composition. Finally, we performed the pair-end 2 150bp sequencing on an illumina Hiseq 4000 platform housed in the LC Sciences. Results:The results showed Genome-wide DNA-methylation analysis also showed differences between the two groups. KEGG analysis revealed that approximately 80% of the differential genes were enriched in metabolic pathways, and significant changes occurred in DNA methylation of genes related to thermogenesis, fatty acid elongation, TCA cycle and oxidative phosphorylation between FT and WT cells。 Conclusions：There are indeed differences between fat-1 transgenic bovine fetal fibroblasts and wild-type in genome-wide DNA methylation detection, and 80% of these differences are related to metabolism.
2021-12-09 | GSE190293 | GEO
Project description:Chromatin immunoprecipitation for the progesterone receptor (PGR) and sequencing was performed on Pgrcre/-mPRALsL/+ mice. These mice exhibit no endogenous PGR expression, yet exhibit constitutive expression of the PGR-A isoform in PGR positive tissues due to the presence of a conditional expression allele. Mice were ovariectomized, rested for two weeks, and treated with progesterone for 1 hour before harvesting and freezing whole uterine tissue.
2019-08-29 | GSE86314 | GEO
Project description:Using a canine custom retinal cDNA microarray our aim is to identify normal gene expression profiling for retina and frontal, occipital and temporal brain cortices Total RNA was isolated using Trizol reagent (Invitrogen, Carlsbad, CA) and further purified by Rneasy mini kit (Qiagen, Valencia, CA). Purity and RNA quality were evaluated by absorbance at 260 nm and by denaturing formaldehyde agarose gel electrophoresis. High quality RNAs with A260/280 ratio over 1.8 and intact 28S and 18S RNA bands were used for microarray analysis. To generate an RNA reference sample for microarray hybridizations, we pooled equal amounts of total RNA from the occipital, temporal, and frontal brain regions collected from three 16-week-old beagles to achieve a homogeneous pool of transcripts. The pooled RNA was divided into aliquots (2μg/μl) and stored at -80 °C until use. For the purpose of this work, four tissue groups have been established, containing five biological replicates for normal retina and three for each respective brain region. After initial validation of reproducibility, only one microarray experiment was used for each sample.
2010-07-01 | E-GEOD-5208 | biostudies-arrayexpress
Project description:To examine the effect of wtAPE1 and ubiquitin-APE1 fusion proteins on global gene expression. Total RNA from HEK293 derivatives that express vector alone (ctl), wt-APE1, or ubiquitin-APE1 fusion in the presence of doxycycline were analyzed. Three HEK293 derivatives expressing none (vector), wtAPE1, or Ub-APE1 fusion linked at APE1's 24th Lys were incubated with/without doxycycline (dox) 2 µg/ml for 16 hr. Each -/+ dox cultures were triplicated. Total RNA was extracted (Qiagen) and then analyzed by LSU's bioinformatics core. The quality and quantity of total RNA was assessed using an ND-1000 Spectrophometer (NanoDrop, Wilmington, DE), NanoChip assay and Bioanalyzer 2100 (Agilent). All samples were of high integrity with RIN # greater than 7 and A260/280 greater than 1.8. Procedures for cDNA synthesis, sense target labeling, and hybridization were carried out as described at http://media.affymetrix.com/support/technical/appnotes/wt_appnote.pdf (Affymetrix, Redwood City, CA). All experiments were performed using GeneChip Exon 1.0 ST Arrays and Whole Transcript Sense Target labeling Assay (version 4, Affymetrix). Overnight hybridization of fragmented single-stranded DNA was carried out in an Affymetrix GeneChip Hybridization Oven 640, then washed and stained with streptavidin-phycoerythrin using GeneChip 450 Microfluidics Station (Affymetrix). Chips were scanned with an Affymetrix High Resolution 3000 (G7) scanner. Signal and background intensities were quantitated by pixel intensity using Affymetrix GeneChip Operating Software (GCOS 1.4). Array quality control assessment was carried out using Robust Multi-chip Analysis (RMA) workflow for Core probesets in Expression Console (Affymetrix). Gene-level analysis for differential gene expression was analyzed in GeneSpring 7.3x (Agilent Technologies) using the sample CHP files. The RMA method was used for the background correction, normalization and average expression measures for the probesets. An expression filter (20th percentile cutoff) was applied to remove genes with low signal intensity values. For differential gene expression analysis, the Welch ANOVA test with asymptotic p-value computation was applied and transcripts with p-value (< 0.05) and fold change (2.0 fold) were selected.
2013-11-23 | GSE25695 | GEO
Project description:Pgrcre/+Foxl2LsL/+ mice showed absence of corpus luteum in the ovary, thus the mice were infertile and continuously at diestrus stage with very thin uterus. Multiple uterine defects were observed. The gland number and penetratioin to the stroma were decreased, and the epithelial stratification and the collagen deposition in the stroma were increased. The whole uterus were collected from 12 weeks old control and mutant mice at diestrus stage for RNA-sequencing. In total, 3515 genes were differentially expressed, with 1746 upregulated and 1769 genes downregulated in the mutant mice. Pathways related to extracellular matrix, gland formation, cell proliferation were altered. The muntant mice transcriptomic changes was significantly postively correlated with the human endometriosis transcrptomic changes.
2020-11-06 | GSE140047 | GEO
Project description:Peripheral blood mononuclear cells were collected from the patients. RNA-seq and analysis was performed on the collected samples. Total RNA was extracted from the tissue using TRIzol® Reagent according the manufacturer’s instructions(Invitrogen) and genomic DNA was removed using DNase I (TaKara). Then RNA quality was determined using 2100 Bioanalyser (Agilent) and quantified using the ND-2000 (NanoDrop Technologies). High-quality RNA sample (OD260/280=1.8~2.2, OD260/230≥2.0, RIN≥6.5, 28S:18S≥1.0, >10μg) is used to construct sequencing library
2024-05-09 | GSE249812 | GEO
Project description:Single cell RNA-seq analysis demonstrated dramatic changes in adult PgrCre/+Srfflox/flox endometrial cell populations compared to Srf flox/flox controls. Uterine Srf ablation caused myeloid immune cell infiltration, and differential expression analysis demonstrated that PgrCre/+Srfflox/flox stromal fibroblasts downregulated progesterone-regulated genes and growth response genes. Proliferating fibroblast populations shifted from G2-M phase-dominant in controls to S-G2 phase-dominant in PgrCre/+Srfflox/flox uteri. PgrCre/+Srfflox/flox epithelial cells overexpressed estrogenic and innate inflammatory genes.
2025-09-30 | GSE294752 | GEO
Project description:To examine the effect of wtAPE1 and ubiquitin-APE1 fusion proteins on global gene expression. Total RNA from HEK293 derivatives that express vector alone (ctl), wt-APE1, or ubiquitin-APE1 fusion in the presence of doxycycline were analyzed. Three HEK293 derivatives expressing none (vector), wtAPE1, or Ub-APE1 fusion linked at APE1's 24th Lys were incubated with/without doxycycline (dox) 2 µg/ml for 16 hr. Each -/+ dox cultures were triplicated. Total RNA was extracted (Qiagen) and then analyzed by LSU's bioinformatics core. The quality and quantity of total RNA was assessed using an ND-1000 Spectrophometer (NanoDrop, Wilmington, DE), NanoChip assay and Bioanalyzer 2100 (Agilent). All samples were of high integrity with RIN # greater than 7 and A260/280 greater than 1.8. Procedures for cDNA synthesis, sense target labeling, and hybridization were carried out as described at http://media.affymetrix.com/support/technical/appnotes/wt_appnote.pdf (Affymetrix, Redwood City, CA). All experiments were performed using GeneChip Exon 1.0 ST Arrays and Whole Transcript Sense Target labeling Assay (version 4, Affymetrix). Overnight hybridization of fragmented single-stranded DNA was carried out in an Affymetrix GeneChip Hybridization Oven 640, then washed and stained with streptavidin-phycoerythrin using GeneChip 450 Microfluidics Station (Affymetrix). Chips were scanned with an Affymetrix High Resolution 3000 (G7) scanner. Signal and background intensities were quantitated by pixel intensity using Affymetrix GeneChip Operating Software (GCOS 1.4). Array quality control assessment was carried out using Robust Multi-chip Analysis (RMA) workflow for Core probesets in Expression Console (Affymetrix). Gene-level analysis for differential gene expression was analyzed in GeneSpring 7.3x (Agilent Technologies) using the sample CHP files. The RMA method was used for the background correction, normalization and average expression measures for the probesets. An expression filter (20th percentile cutoff) was applied to remove genes with low signal intensity values. For differential gene expression analysis, the Welch ANOVA test with asymptotic p-value computation was applied and transcripts with p-value (< 0.05) and fold change (2.0 fold) were selected. Three cell lines (HEK293/pcDNA-FRT empty vector, HEK293/pcDNA-FRT wtAPE1, HEK293/pcDNA-FRT ubAPE1), +/- 2µg/ml doxycycline for 16 h. Triplicates of each cell line/condition. Replicates were averaged.
2013-11-23 | E-GEOD-25695 | biostudies-arrayexpress
Project description:Simple steatosis (SS) and non-alcoholic steatohepatitis (NASH) are subtypes of non-alcoholic fatty liver disease. The difference in pathogenesis between SS and NASH is still not clear. MicroRNAs (miRNAs) are endogenous, non-coding short RNAs that regulate gene expression. The aim of this study was to examine the relationship of miRNA expression profiles with SS and NASH in animal models and humans. Animal models DD Shionogi (DS), Fatty Liver Shionogi (FLS Wild; W), and FLS ob/ob mice were subjected as the normal control, SS model and NASH model, respectively. Male DS mice (Riken BRC No. 03706) were provided by Riken bio-resource center through the national bio-resource project of Japan. Male FLS W and male FLS-ob/ob mice were obtained from Shionogi research laboratories (Shiga, Japan). Animals were housed in a room maintained at a controlled temperature of 24 ± 2 Ë?C under a 12-h light-dark cycle. Animals were provided ad libitum access to water and standard pellet feed. Five male mice of every strain (24 weeks old; mean body weight of DS, FLS W and FLS-ob/ob were 31.1 ± 1.1g, 38.4 ± 3.4g and 56.0 ± 5.0g, respectively) were sacrificed under pentobarbital anesthesia by whole blood collection from the right ventricle. The livers were cut into about 200 mg pieces and fixed in 10% formalin for histological analysis or fresh-frozen in liquid nitrogen and stored at -80 °C in a freezer until use. miRNA expression analysis Total RNAs in mouse liver were isolated using TrizolReagent (Life Technologies, California, USA) as described the manufacturerâ??s protocol. The quality of total RNA samples was checked by applying the RNA Integrity Number (RIN) which is calculated by a proprietary algorithm of the Agilent 2100 Bioanalyzer expert software (Agilent technologies, CA, USA). Only high quality RNA, with RNA integrity number (RIN) greater than 8 and A260/280 and A260/230 greater than 1.8, will be considered for microarray analysis. TaqMan® Array Rodent MicroRNA A Card v2.0 (Thermo Fisher Scientific, MA, USA) was used to assess 375 miRNA expression profiles in mouse liver tissue.
2016-07-03 | E-GEOD-69670 | biostudies-arrayexpress
Project description:Super-Low Input Carrier Cap Analysis of Gene Expression (SLIC-CAGE) to identify transcription start sites (TSS) and existing genome-wide maps of islet histone marks to characterise the contribution of transcriptional regulation of LKB1-mediated control of gene expression in mouse β-cells. SLIC-CAGE was performed as in (Cvetesic et al. - doi: 10.1101/gr.235937.118) using 100 ng of total RNA extracted as described above from islets isolated from a 18 week old female mouse on a C57BL6/J background. 12 cycles were performed for library amplification. The amplified library was purified with AMPure XP beads and visualised using a HS DNA chip (Bioanalyzer, Agilent). The library was sequenced on a HiSeq2500 instrument with paired-end, 150bp reads). SLIC-CAGE paired-end sequencing data was aligned to GENCODE assembly annotation version GRCm38.p6 using the STAR alignment tool v2.4.2a.
2024-09-19 | E-MTAB-14336 | biostudies-arrayexpress