Project description:PURPOSE: To provide a detailed gene expression profile of the normal postnatal mouse cornea. METHODS: Serial analysis of gene expression (SAGE) was performed on postnatal day (PN)9 and adult mouse (6 week) total corneas. The expression of selected genes was analyzed by in situ hybridization. RESULTS: A total of 64,272 PN9 and 62,206 adult tags were sequenced. Mouse corneal transcriptomes are composed of at least 19,544 and 18,509 unique mRNAs, respectively. One third of the unique tags were expressed at both stages, whereas a third was identified exclusively in PN9 or adult corneas. Three hundred thirty-four PN9 and 339 adult tags were enriched more than fivefold over other published nonocular libraries. Abundant transcripts were associated with metabolic functions, redox activities, and barrier integrity. Three members of the Ly-6/uPAR family whose functions are unknown in the cornea constitute more than 1% of the total mRNA. Aquaporin 5, epithelial membrane protein and glutathione-S-transferase (GST) omega-1, and GST alpha-4 mRNAs were preferentially expressed in distinct corneal epithelial layers, providing new markers for stratification. More than 200 tags were differentially expressed, of which 25 mediate transcription. CONCLUSIONS: In addition to providing a detailed profile of expressed genes in the PN9 and mature mouse cornea, the present SAGE data demonstrate dynamic changes in gene expression after eye opening and provide new probes for exploring corneal epithelial cell stratification, development, and function and for exploring the intricate relationship between programmed and environmentally induced gene expression in the cornea. Keywords: other
Project description:A transcriptome study in mouse hematopoietic stem cells was performed using a sensitive SAGE method, in an attempt to detect medium and low abundant transcripts expressed in these cells. Among a total of 31,380 unique transcript, 17,326 (55%) known genes were detected, 14,054 (45%) low-copy transcripts that have no matches to currently known genes. 3,899 (23%) were alternatively spliced transcripts of the known genes and 3,754 (22%) represent anti-sense transcripts from known genes. Mouse hematopoietic stem cells were purified from bone marrow cells using negative and positive selection with a Magnetic-Activated Cell Sorter (MACS). total RNA and mRNA were purified from the purified cells using Trizol reagent and magnetic oligo dT beads. Double strand cDNAs were synthesized using a cDNA synthesis kit and anchored oligo dT primers. After NlaIII digestion, 3’ cDNAs were isolated and amplified through 16-cycle PCR. SAGE tags were released from the 3’ cDNA after linker ligation. Ditags were formed, concatemerized and cloned into a pZERO vector. Sequencing reactions were performed with the ET sequencing terminator kit. Sequences were collected using a Megabase 1000 sequencer. SAGE tag sequences were extracted using SAGE 2000 software.
Project description:A transcriptome study in mouse hematopoietic stem cells was performed using a sensitive SAGE method, in an attempt to detect medium and low abundant transcripts expressed in these cells. Among a total of 31,380 unique transcript, 17,326 (55%) known genes were detected, 14,054 (45%) low-copy transcripts that have no matches to currently known genes. 3,899 (23%) were alternatively spliced transcripts of the known genes and 3,754 (22%) represent anti-sense transcripts from known genes.
Project description:Transcription factor 21 (TCF21) is a basic helix-loop-helix protein required for developmental specification of cardiac fibroblasts from epicardial progenitor cells that normally surround and invade the heart. In the adult heart, TCF21 is expressed in tissue-resident fibroblasts but is downregulated in response to injury or stimuli leading to myofibroblast differentiation. These findings led to the hypothesis that Tcf21 could be a regulator of fibroblast cell fate in the adult mammalian heart and contribute to cardiac fibrosis. Here, single-cell RNA-sequencing was used to study the effect of loss of Tcf21 in cardiac fibroblasts in adult mouse hearts at baseline and after an ischemic cardiac injury (myocardial infarction).
Project description:SILAC based protein correlation profiling using size exclusion of protein complexes derived from Mus musculus tissues (Heart, Liver, Lung, Kidney, Skeletal Muscle, Thymus)
Project description:SILAC based protein correlation profiling using size exclusion of protein complexes derived from seven Mus musculus tissues (Heart, Brain, Liver, Lung, Kidney, Skeletal Muscle, Thymus)
Project description:We created mice, which are deficient for Myc specifically in cardiac myocytes by crossing crossed Myc-floxed mice (Mycfl/fl) and MLC-2VCre/+ mice. Serial analysis of earlier stages of gestation revealed that Myc-deficient mice died prematurely at E13.5-14.5. Morphological analyses of E13.5 Myc-null embryos showed normal ventricular size and structure; however, decreased cardiac myocyte proliferation and increased apoptosis was observed. BrdU incorporation rates were also decreased significantly in Myc-null myocardium. Myc-null mice displayed a 3.67-fold increase in apoptotic cardiomyocytes by TUNEL assay. We examined global gene expression using oligonucleotide microarrays. Numerous genes involved in mitochondrial death pathways were dysregulated including Bnip3L and Birc2. Hearts were taken from wide type and Myc-null Mouse embryos at E13.5 under the dissecting scope. Cardiac myocyte RNA was isolated using TRIZOL®Reagent Total RNA (100 ng) was hybridized to the Sentrix® MouseRef-8 Expression BeadChip that contains probes for ~24,000 transcripts. GeneChips were scanned using the Hewlett-Packard GeneArray Scanner G2500A. The data were analyzed with Illumina Inc. BeadStudio version 1.5.0.34 and normalized by rank invariant method.