Project description:Differentiation of hemopoietic stem cells into granulocytes is characterized by distinct changes in the transcriptome. We analyzed mRNA expression in primary murine myeloid cells at four successive stages of hemopoietic differentiation; Lin- Sca1+ cKit+ stem/progenitor cells (LSKs), promyelocytes, myelocytes and granulocytes.
Project description:Differentiation of hemopoietic stem cells into granulocytes is characterized by distinct changes in the transcriptome. We analyzed mRNA expression in primary murine myeloid cells at four successive stages of hemopoietic differentiation; Lin- Sca1+ cKit+ stem/progenitor cells (LSKs), promyelocytes, myelocytes and granulocytes. Using fluorescence–activated cell sorting, we isolated primary murine myeloid cells at four successive stages of hemopoietic differentiation; Lin- Sca1+ cKit+ stem/progenitor cells (LSK), promyelocytes, myelocytes and granulocytes.
Project description:This study sought to determine the dynamic changes of miRNA expression during mouse granulopoiesis. We not only performed analyses of miRNA expression levels in whole cells but also analyzed purified nuclear and cytoplasmic cell fractions to profile miRNA subcellular localization. qRT-PCR analysis of miRNAs was performed on whole cell, nuclear and cytoplasmic RNAs extracted from mouse hemopoietic stem cells (LSKs), promyelocytes, myelocytes and granulocytes. 100 ng of RNA was reversed transcribed using the Taqman miRNA Reverse Transcription Kit and Megaplex RT Primers rodent pool A and B (Life Technologies). Complementary DNA (cDNA) was amplified using a TaqMan rodent microRNA A and B Array v2.0 (Life Technologies) with TaqMan Universal PCR Master Mix on an ABI 7900HT Sequence Detection System.
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:To investigate the differences in microRNA expression profiles between fibrotic and normal livers, we performed microRNA microarrays for total RNA extracts isolated from mouse livers treated with carbontetrachloride (CCl4) or corn-oil for 10 weeks (n=3/group). MicroRNAs were considered to have significant differences in expression level when the expression difference showed more than two-fold change between the experimental and control groups at p<0.05. We found that 12 miRNAs were differentially expressed in CCl4-induced fibrotic liver.
Project description:Using mRNA-seq, we determined intron retaining genes that were differentially regulated in FACS purified cells at three progressive stages of mouse granulopoiesis; CD34+Kit+Gr-1low promyelocytes, CD34-Kit-Gr-1mid myelocytes and CD34-Kit-Gr-1high granulocytes. We found that IR affects 86 genes, including those specific to granulocyte (Lyz2 and MMP8) and nuclear architecture (Lmnb1 and Lbr). IR was associated with the decrease in protein levels measured by mass spectrometry (P=0.0015, binomial test). There was a significant overlap of IR between human and mouse (P=2.85E-22, hypergeometric test), showing that IR is conserved.Inhibition of NMD in granulocytes resulted in marked accumulation of 39/86 intron retaining mRNAs (P<0.05, RUV procedure with Holm-Bonferroni correction), indicating that IR triggers NMD to downregulate mRNA and protein expression. Sequencing of polyadenylated RNA from three types of myeloid cells (promyelocytes, myelocytes and granulocytes) using Illumina GAIIx