RNAi knock down of Klf4 in mouse embryonic stem cells
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ABSTRACT:
ORGANISM(S): Mus musculus
SUBMITTER: Yong Qian
PROVIDER: E-MNIA-92 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:To characterize Klf4 downstream genes, we performed microarray analyses of the ES cells knocked down by siRNA of Klf4 gene. Keywords: individual genetic characteristic design ES cells grow without feeders on gelatin coated in complete ES medium. Cells were incubated at 37 0C; 5% CO2. ES cells were transfected with siRNA and pCAGEGFP-IP plasmid. After transfection, cells were cultured for 24 hours, following to drag selection for more 24 hours in the presence of puromycin to ensure concentration of transfectants. Total RNA were extracted with Trizol reagent (Sigma) according to the manufacturerâ??s instructions.
2010-06-10 | E-GEOD-5930 | biostudies-arrayexpress
Project description:Thyroid hormone (TH) controls the remodeling of the pancreas and the liver. TH-induces dedifferentiation of the exocrine pancreas to a progenitor state (Proc. Nat. Acad Sci. 105, 8962-8967 (2008)) and it remodels the endocrine pancreas (Dev. Biol. 328, 384-391 (2009)). The redifferentiated frog pancreas resembles closely the pancreas of other typical vertebrates. Two pancreas arrays were carried out. The first one studied gene expression changes at different developmental stages of Xenopus laevis during metamorphosis. The second array studies gene expression changes at varying times after the addition of TH to premetamorphic tadpoles. Keywords: co-expression design,development or differentiation design,reference design,time series design Overall design: Thyroid hormone (TH) controls remodeling of the liver. The microarray was carried out to identify changes in gene expression at different stages of development during metamorphosis. This is part 2, done on the 22K Xenopus platform version 1. This dataset was from livers of Xenopus tadpoles (NF52, NF62, and NF66). Each was made in triplicate. Samples in all 3 parts of the study received the same thyroid hormone levels.
2010-05-16 | E-GEOD-16017 | biostudies-arrayexpress
Project description:Thyroid hormone (TH) controls the remodeling of the pancreas and the liver. TH-induces dedifferentiation of the exocrine pancreas to a progenitor state (Proc. Nat. Acad Sci. 105, 8962-8967 (2008)) and it remodels the endocrine pancreas (Dev. Biol. 328, 384-391 (2009)). The redifferentiated frog pancreas resembles closely the pancreas of other typical vertebrates. Two pancreas arrays were carried out. The first one studied gene expression changes at different developmental stages of Xenopus laevis during metamorphosis. The second array studies gene expression changes at varying times after the addition of TH to premetamorphic tadpoles. Keywords: co-expression design,development or differentiation design,reference design,time series design Overall design: Thyroid hormone (TH) controls remodeling of the pancreas. The microarray was carried out to identify changes in gene expression at different stages of development during metamorphosis.. This is part 3, done on the 22K Xenopus platform version 2. This dataset was from pancreas of Xenopus tadpoles (NF52, NF62, and NF66). Each was made in triplicate. Samples in all 3 parts of the study received the same thyroid hormone levels.
2010-05-16 | E-GEOD-16018 | biostudies-arrayexpress
Project description:We present a microarray experiment with the same tet-inducible system but with multiple time points within 24 hr to capture early responses to Oct4 suppression. These data were analyzed together with published genome-wide ChIP data. We found that most tentative target genes (TTG) of Oct4 in ES cells were activated by Oct4 and only a few genes were suppressed. The same method was then applied to find target genes of Sox2 and Nanog based mostly on previously published data. Because the interaction between Oct4, Sox2, and Nanog is supported by immunoprecipitation, functional analysis, and co-localization of binding sites, we explored the relationships between their target genes. Keywords: time series design We used ZHBTc4 ES cells with a Tet-inducible Oct4 transgene. Cells were cultured for 2 passages on gelatin-coated plates in order to remove feeder cells and then transferred to gelatin-coated 6-well plates at the density of 1-2x105 cells/well and cultured in complete ES medium. Tetrocycline (TC) was added at 24 hr after cell plating, and then cells were harvested at 0hr (before adding TC), 3 hr, 6 hr, 12 hr, and 24 hr (2 replications each). RNA samples for later time points (24, 48, 72, 96, and 120 hr) were obtained from our earlier experiment with 3 replications. Two Nanog over-expressing clones were tested: (1) integrated transgene in MG1.19 parental cell line and (2) episomal transgene in EBRTcH3 parental line. EBRTcH3 and MG1.19 ESC lines were kind gifts of Dr. Hitoshi Niwa (RIKEN Center for Developmental Biology, Kobe, Japan). Both transgenic and parental cell lines were cultured for 2 passages on gelatin-coated plates and then transferred to gelatin-coated 6-well plates at the density of 1-2x105 cells/well and cultured for 3 days in 3 different conditions: (1) complete ES medium (see above); (2) complete medium without LIF, and (3) complete medium with 1 uM RA. Cells were cultured at 37 0C and 5% CO2 condition and the culture medium was changed daily. Over-expression of Nanog was evaluated by PCR. Total RNAs were extracted using TrizolTM (Invitrogen, USA) and Phase lock gelTM columns (Eppendorf/Brinkman) according to the manufacturer???s protocol. Total RNAs were precipitated with isopropanol, washed with 70% ethanol, and dissolved in DEPC-treated H2O. 2.5 ug of total RNA samples were labeled with Cy3-CTP using a Low RNA Input Fluorescent Linear Amplification Kit (Agilent, USA). A reference target (Cy5-CTP-labeled) was prepared from the Universal Mouse Reference (UMR) RNA (Stratagene, USA).
2008-04-24 | E-GEOD-8617 | biostudies-arrayexpress
Project description:E12.5 mouse whole embryo and E12.5 placenta total RNA were pooled to create 25:75, 50:50, and 75:25 ratio mixtures, based on Bioanalyzer quantitation. These samples, along with the original unmixed RNAs, were used as templates for duplicate linear amplification labeling reactions. cRNA target mixtures were hybridized against a Universal Mouse Reference (Stratagene). Pairwise comparison using the NIA Microarray Analysis (ANOVA) software produced log ratios, which were compared to the expected log ratios for genes showing statistically significant (FDR<0.05) differential expression between unmixed embryo and placenta.
2005-01-01 | E-MNIA-67 | biostudies-arrayexpress
Project description:E12.5 mouse whole embryo, E12.5 placenta, embryonic stem (ES) cells, and trophoblast stem (TS) cells were compared on a novel microarray design to validate the system. The in situ-synthesized 60-mer oligonucleotide microarray platform contains approximately 44,000 DNA features, and was designed to detect transcripts from all known genes, as well as about 5,000 potential genes, as identified by the NIA Mouse Gene Index 2.0. Seven external RNA controls were derived from intronic and intergenic yeast genome sequences, and 60-mer probes were spotted 10 time each on the slides. These spike-in RNA controls were diluted to cover 7 orders of magnitude in initial concentration, and used to define a function relating signal intensity to transcript abundance.
2005-01-01 | E-MNIA-66 | biostudies-arrayexpress
Project description:To characterize the differentiation by Sox2 KO, we performed microarray analyses of mouse ES cell line 2TS22C during the time-course being induced of Sox2 KO Inducible Sox2 KO cell line 2TS22C were assigned to eight experimental groups and cultured in GMEM medium (SIGMA) with LIF at 37°C in an atmosphere of 5% CO2 until the specified time point before the harvest: (1)for controls, cultured in the absence of Tc; (2) for 3h after the administration of Tc; (3) for 6h; (4) for 12h; (5) for 24h; (6) for 48h; (7) for 72h; (8) for 96h.
2010-06-11 | E-GEOD-5895 | biostudies-arrayexpress
Project description:Thyroid hormone (TH) controls the remodeling of the pancreas and the liver. TH-induces dedifferentiation of the exocrine pancreas to a progenitor state (Proc. Nat. Acad Sci. 105, 8962-8967 (2008)) and it remodels the endocrine pancreas (Dev. Biol. 328, 384-391 (2009)). The redifferentiated frog pancreas resembles closely the pancreas of other typical vertebrates. Two pancreas arrays were carried out. The first one studied gene expression changes at different developmental stages of Xenopus laevis during metamorphosis. The second array studies gene expression changes at varying times after the addition of TH to premetamorphic tadpoles. Keywords: cell cycle design,co-expression design,reference design,time series design Thyroid hormone (TH) controls remodeling of the pancreas. The micro array was carried out to identify changes in gene expression between the tadpole and frog pancreas. This is part 1, done on the 44K Xenopus platform. This dataset was used only for one pancreas experiment with several data points (control, 12h, 24h, 48h and frog). Each was made in triplicate. Samples in all 3 parts of the study received the same thyroid hormone levels.
2010-05-16 | E-GEOD-16074 | biostudies-arrayexpress
Project description:E12.5 mouse whole embryo and E12.5 placenta total RNA were pooled to create 25:75, 50:50, and 75:25 ratio mixtures, based on Bioanalyzer quantitation. These samples, along with the original unmixed RNAs, were used as templates for duplicate linear amplification labeling reactions. cRNA target mixtures were hybridized against a Universal Mouse Reference (Stratagene). Pairwise comparison using the NIA Microarray Analysis (ANOVA) software produced log ratios, which were compared to the expected log ratios for genes showing statistically significant (FDR<0.05) differential expression between unmixed embryo and placenta. Keywords: cell type comparison design,development or differentiation design,normalization testing design,reference design E12.5 mouse whole embryo and E12.5 placenta total RNA were pooled to create 25:75, 50:50, and 75:25 ratio mixtures, based on Bioanalyzer quantitation. These samples, along with the original unmixed RNAs, were used as templates for duplicate linear amplification labeling reactions. cRNA target mixtures were hybridized against a Universal Mouse Reference (Stratagene). Pairwise comparison using the NIA Microarray Analysis (ANOVA) software produced log ratios, which were compared to the expected log ratios for genes showing statistically significant (FDR<0.05) differential expression between unmixed embryo and placenta.
2010-06-09 | E-GEOD-3508 | biostudies-arrayexpress
Project description:This SuperSeries is composed of the following subset Series: GSE16017: Thyroid Hormone (TH) Controls The Remodeling Of The Pancreas And The Liver, Part B GSE16018: Thyroid Hormone (TH) Controls The Remodeling Of The Pancreas And The Liver, Part C GSE16074: Thyroid Hormone (TH) Controls The Remodeling Of The Pancreas And The Liver, Part A Refer to individual Series
2010-05-16 | E-GEOD-16075 | biostudies-arrayexpress