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Expression data from mammary epithelial cells harvested from 8-9 week old C3H female mice - various isolation conditions

ABSTRACT: Numerous lineage analysis studies have used FACS sorting as a separation technique prior to measuring mRNA expression patterns. To assess if FACS sorting causes non-specific changes in gene expression, we collected a heterogeneous population of mammary epithelial cells from three biological replicates. We then performed gene expression analysis on each replicate either prior to trypsin digestion, immediately following trypsin digestion, or following both the trypsin digestion and a mock FACS sort. In this dataset, we include gene expression data obtained from isolated murine mammary epithelial cells either prior to trypsin digestion, immediately following trypsin digestion, or following the trypsin digestion and a mock FACS sort. Nine total samples were analyzed (3 biological replicates of three experimental conditions). Using LIMMA packages gene-wise comparisons were made between untrypsinized and trypsinized replicates as well as between trypsinized and mock FACS sorted replicates using and Pâ¤0.05 and a â¥1.5-fold difference between conditions.

ORGANISM(S): Mus musculus

SUBMITTER: Ian Macara

PROVIDER: E-GEOD-61167 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Expression data from mammary epithelial cells harvested from 8-9 week old C3H female mice - various isolation conditions

Project description:Numerous lineage analysis studies have used FACS sorting as a separation technique prior to measuring mRNA expression patterns. To assess if FACS sorting causes non-specific changes in gene expression, we collected a heterogeneous population of mammary epithelial cells from three biological replicates. We then performed gene expression analysis on each replicate either prior to trypsin digestion, immediately following trypsin digestion, or following both the trypsin digestion and a mock FACS sort. In this dataset, we include gene expression data obtained from isolated murine mammary epithelial cells either prior to trypsin digestion, immediately following trypsin digestion, or following the trypsin digestion and a mock FACS sort.

2014-09-05 | GSE61167 | GEO

Transcription profiling of mouse thymic stromal cells from 6-12 week old wild type (C57BL/6) and Aire knock-outs (mixed background)

Project description:Gene expression in different thymic stromal cells and subsets thereof was analyzed in 6-12 week old wild type (C57BL/6) and Aire knock-out (mixed background) mice. Thymic stromal cells were purified by sequential enzymatic digestion (collagenase, collagenase/dispase and trypsin) followed by gradient centrifugation and FACS sorting. Sort criteria were as follows: dendritic cells (CD11c+, F4/80 -), macrophages (F4/80+, CD11c-), cTECs (CD45â/lo, CDR1/Ly51+, Ep-CAM+) and mTECs (CD45â/lo, CDR1/Ly51â, Ep-CAM+). mTECs of wild-type and Aire knock-out mice were further subdivided according to CD80 expression levels.,For microarray analysis total RNA from thymic stromal cell samples of two independent experiments was pre-amplified and biotinylated by two rounds of cDNA synthesis and in vitro transcription. Fluorescence readings were evaluated by using Microarray Suite 5.0 software.

2007-12-15 | E-GEOD-2585 | biostudies-arrayexpress

promiscuous gene expression in the mouse thymus

Project description:Gene expression in different thymic stromal cells and subsets thereof was analyzed in 6-12 week old wild type (C57BL/6) and Aire knock-out (mixed background) mice. Thymic stromal cells were purified by sequential enzymatic digestion (collagenase, collagenase/dispase and trypsin) followed by gradient centrifugation and FACS sorting. Sort criteria were as follows: dendritic cells (CD11c+, F4/80 -), macrophages (F4/80+, CD11c-), cTECs (CD45–/lo, CDR1/Ly51+, Ep-CAM+) and mTECs (CD45–/lo, CDR1/Ly51–, Ep-CAM+). mTECs of wild-type and Aire knock-out mice were further subdivided according to CD80 expression levels. For microarray analysis total RNA from thymic stromal cell samples of two independent experiments was pre-amplified and biotinylated by two rounds of cDNA synthesis and in vitro transcription. Fluorescence readings were evaluated by using Microarray Suite 5.0 software. Keywords = thymus Keywords = promiscuous gene expression Keywords = AIRE Keywords: ordered

2005-04-28 | GSE2585 | GEO

mouse dendritic cell subtypes proteomics

Project description:dendritic cells = improtant APC -different subtypes characterized by surface markers by FACS -combination of FACS and LC-MS powerful tool, especially for low cell numbers -direct sorting of cells into SP3-compatible lysis buffer -new protocol without washing steps after sort and before protein digestion

2024-07-25 | PXD031960 | JPOST Repository

RNA-seq of human colorectal cancer HCT116 and HT29 cells by the distinct conditions of trypsin digestion

Project description:In the process of cell culture in vitro, we usually find that the cells show signs of aging, which is that the passed cells are unable to stick to the wall. Experience tells us that the time it takes for trypsin to digest the adherent cells is important. If the digestion time is too short, enough cells cannot be obtained for passage, while if the digestion time is too long, a large number of senescent cells will also be digested. The substances released by the death and decomposition of these senescent cells will affect the function of those active cells, resulting in the failure of the recovery of the whole daughter cell line. This project starts from the question of the global difference among cells undergoing the multiple rounds of mitosis in one culture dish. RNA sequencing was conducted to investigate the transcriptome signatures of cells with different sensitivity response to trypsin.

2024-12-21 | E-MTAB-14651 | biostudies-arrayexpress

Gene expression profiles of the adult mesangial cell (GUDMAP Series_id: 37)

Project description:The long term objective is to create an encyclopedia of the expression levels of all genes in multiple components of the developing kidney. The central thesis is straightforward. The combination of fluorescent activated cell sorting (FACS) plus microarray analysis offers a powerful, efficient and effective method for the creation of a global gene expression atlas of the developing kidney. Microarrays with essentially complete genome coverage can be used to quantitate expression levels of every gene in FACS isolated components of the developing kidney. The ensuing rapid read-out provides an expression atlas that is more sensitive, more economical and more complete than would be possible by in situ hybridizations alone. Meis1-GFP BAC transgenic mice were utilized to isolate the mesangial cells from adult kidneys. The mesangial cells were isolated from the glomerulus using collagenase digestion, differential sieving, trypsin treatment and FACS. The RNA was isolated from purified mesangial cells and the gene expression profiles were determined by microarrays.

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Transcriptional profiling of embryonic skeletal muscle stem/progenitor cells

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2013-11-08 | E-GEOD-52192 | biostudies-arrayexpress

Expression data from osteocytes in virgin,lactating and post lactation mice

Project description:We have previously shown that during lactation, osteocytes directly remodel their perilacunar and pericanalicular matrix, thereby mobilizing calcium and contributing to maternal bone loss. To identify genes potentially responsible for perilacunar remodeling, microarray analysis was performed on osteocytes from CD-1 virgin and lactating mice and mice sacrificed on day 7 post weaning. By comparing how gene expression changes in osteocytes among virgin, lactation and post lactation, the goal is to identify the genes osteocytes use to remodel their perilacunar matrix To identify genes responsible for perilacunar remodeling, microarray analysis was performed on tibiae from CD-1 virgin and lactating mice and mice sacrificed on day 7 post weaning. Periosteum and bone marrow were removed from the tibiae and the bone fragments underwent sequential collagenase/trypsin digestion and were examined histologically to ensure removal of all surface cells and confirm osteocyte enrichment prior to RNA extraction. Microarray Affy M430 arrays, n=3 per group were performed and Genepattern and Bioconductor were used to analyse the microdata.

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Promiscuous gene expression in human medullary thymic epithelial cell subsets

Project description:Different human mTEC subsets (MUC1, CEACAM5 and SGLT1) were purified by sequential enzymatic digestion (collagenase/dispase, trypsin) followed by enrichment using magnetic beads (CD45 beads, Miltenyi Biotech) and FACS sorting. Cells of the surface phenotype CD45-, CDR2-, EpCAM+ were further subdivided into MUC1+/MUC1-, CEACAM5+/CEACAM5- and SGLT1+/SGLT1- fractions. RNA was isolated using μMACS™ SuperAmp™ protocol (Miltenyi Biotec) and hybridized to Illumina Whole-Genome Expression Beadchips. Gene expression of Antigen-positive and Antigen-negative mTEC subsets was compared.

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