Lifeact-EGFP mouse
Ontology highlight
ABSTRACT:
PROVIDER: PRJEB33979 | ENA |
REPOSITORIES: ENA
Similar Datasets
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2011-02-16 | E-MTAB-537 | biostudies-arrayexpress
Project description:Yeast enolase (Eno2p) conjugated with EGFP and Flag-tag (Eno2p-EGFP-FLAG) and Eno2p with V22A substitution (Eno2V22Ap) conjugated with EGFP and Flag-tag (Eno2V22Ap-EGFP-FLAG) were produced in baker's yeast S. cerevisiae. After semi-anaerobic culture at 30 ˚C for 12h, cells producing Eno2p-EGFP-FLAG formed fluorescent foci, while cells producing Eno2V22Ap-EGFP-FLAG did not. The cells were collected and lysed, and proteins Eno2p-EGFP-FLAG or Eno2V22Ap-EGFP-FLAG and the associated proteins were coimmunoprecipitated using ANTI-FLAG M2 affinity gel and analyzed. Data contain two biological replicates and two technical replicates (n = 4). As the results, 96 proteins were detected with both recombinant Eno2p-EGFP-FLAG and Eno2V22Ap-EGFP-FLAG protein, 29 proteins were detected only with recombinant Eno2p-EGFP-FLAG protein, and 16 proteins were detected only with recombinant Eno2V22Ap-EGFP-FLAG protein. Data Processing/Data Analysis: The separated analytes were detected on an LTQ Velos linear ion trap mass spectrometer (Thermo Scientific). For data-dependent acquisition, the method was set to automatically analyze the five most intense ions observed in the MS scan. The mass spectrometry data were used for protein identification by the Mascot search engine on Protein Discoverer software (ver. 1.2, Thermo Scientific) against the information in the Saccharomyces Genome Database (SGD; http://www.yeastgenome.org). Search parameters for peptide identification included a precursor mass tolerance of 1.2 Da, a fragment mass tolerance of 0.8 Da, a minimum of one tryptic terminus, and a maximum of one internal trypsin cleavage site. Cysteine carbamidomethylation (+57.021 Da) and methionine oxidation (+15.995 Da) were set as a differential amino acid modification. The data were then filtered at a q value ≤ 0.01 corresponding to 1% FDR at the spectral level.
2014-02-25 | PXD000173 | Pride
Project description:Mapttm1(EGFP)Klt/J mice (Mapt-EGFP; The Jackson Laboratory, Bar Harbor, ME, USA; stock 004779) carry a knock-in of the EGFP coding sequence in the first exon of the microtubule-associated protein tau (Mapt) gene producing a cytoplasmic EGFP fused to the first 31 amino acids of MAPT. EGFP expression marks neurons including enteric neurons regardless of their lineage, closely patterning the expression of neuron-specific beta-tubulin III (TUBB3). Mapt-EGFP ice were backcrossed to C57BL/6J (Jackson Laboratory strain #:000664) for three to five generations at Mayo Clinic. Six male and six female Mapt-EGFP mice (54-98 days of age) underwent surgical laparotomy in 3 groups (surgery #1: 1 male and 1 female, surgery #2: 3 males and 1 female, surgery #3: 2 males and 4 females) under pentobarbital (50mg/kg) anesthesia. The celiac ganglion of each mouse was injected with 3-5 μL of 25 mg/mL Alexa Fluor 647-labeled cholera toxin subunit B (CTB-AF647; Thermo Fisher Scientific, Waltham, MA, USA) with the intention of labeling the cell soma of intestinofugal neurons in the myenteric plexus of the colon. The animals were killed 3-4 days after surgery. The muscularis externa of the colon from each Mapt-EGFP mouse was pooled together between all mice of the same surgery date (2, 4, and 6 mice) and mechanically and enzymatically dissociated into single cells with a two-step process that first enriches for cells within myenteric ganglia (PMCID: PMC8114175). The pooled cells from each group of mice formed one biological replicate and subjected to FACS immediately after dissociation to generate populations of Mapt-EGFP+ neurons with or without the CTB-AF647 tracer and Mapt-EGFP− non-neuronal cells. The frequency of Mapt-EGFP+CTB-AF647+ neurons was approximately 125-fold lower than that of Mapt-EGFP+CTB-AF647− neurons and RNA from these preparations did not pass quality control. Therefore, only data from Mapt-EGFP+CTB-AF647− neurons were analyzed and referred to as Mapt-EGFP+ cells. Total RNA was isolated from Mapt-EGFP+ colonic neurons and Mapt-EGFP− myenteric cells using RNA-Bee (AMSBIO, Cambridge, MA, USA) and purified with RNeasy Mini Kit (Qiagen, Germantown, MD, USA). RNA quality was tested using Agilent Electropherogram (Agilent Technologies, Santa Clara, CA, USA) and hybridized to Affymetrix Mouse Genome 430.2 gene expression microarrays (Thermo Fisher Scientific, Waltham, MA, USA). This study utilized Affymetrix Mouse Genome 430.2 oligonucleotide microarray analysis to charaterize the transcriptome of Mapt-EGFP+ neurons and Mapt-EGFP- non-neuronal myenteric cells isolated from the colon of Mapttm1(EGFP)Klt/J mice.
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Project description:Gene expression profiles of ZHBTc4 ES cells expressing EGFP, Oct4-EGFP, Nr5a2-EGFP under CAG promoter. Monoclonal cell lines selected by Puromycin were used for analysis. Each of the cell lines was cultured in doxycycline containing media for endogenous Oct4 knock-down. Pupose of this experiment is to investigate the possibility that forced expression of Nr5a2 can replace Oct4 function in the self-renewal of ES cells. Monoclonal ZHBTc4 ES cells expressing EGFP vs Nr5a2-EGFP, Oct4-EGFP vs Nr5a2-EGFP, no replication
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