Project description:To better understand the molecular bases of the oncogenic cooperation between trisomy 21 and RAS/MAPK pathway activation, KRASG12D was overexpressed in wild-type (WT) or partially trisomic (Ts1Rhr) bone marrow cells. Transduced mCherry-positive murine B cell progenitors were platted in methylcellulose to form B cell precursors colonies (CFU-preB). After a week, B220+/CD19+/CD25- B cells were harvested to perform RNA-sequencing experiments.
Project description:Hypothalamic neuronal populations are central regulators of energy homeostasis and reproductive function. However, the ontogeny of these critical hypothalamic neuronal populations is largely unknown. Here, we reveal novel cellular fates of the hypothalamic Pomc-expressing precursors by combining mouse genetics with a conditional viral ribosome-tagging approach to phenotype neurons. Our results show that the Pomc-expressing precursors differentiate into discrete neuronal subpopulations that mediate not only energy balance (POMC and AgRP) but also reproductive physiology (Kisspeptin).
Project description:Hypothalamic neuronal populations are central regulators of energy homeostasis and reproductive function. However, the ontogeny of these critical hypothalamic neuronal populations is largely unknown. Here, we reveal novel cellular fates of the hypothalamic Pomc-expressing precursors by combining mouse genetics with a conditional viral ribosome-tagging approach to phenotype neurons. Our results show that the Pomc-expressing precursors differentiate into discrete neuronal subpopulations that mediate not only energy balance (POMC and AgRP) but also reproductive physiology (Kisspeptin). Punches containing the mediobasal hypothalamus of Pomc-Cre:RiboTag mice and Pomc-Cre mice injected with the RiboTag viral vector (AAV-DIO-RiboTag) were homogenized and incubated with anti-hemagglutin (HA) antibodies and protein A/G magnetic beads to isolate the polysome-associated mRNAS in the Pomc-derived lineage and in adult POMC neurons, respectively. To identify differentially expressed transcripts, RNA from the immunoprecipitates was extracted, amplified, labelled and hybridized to MouseRef-8 v2 expression beadchips.
Project description:To analyze gene expression in in regulatory T cell precursors that develop in the absence of a functional Foxp3 protein as compared to that of normal regulatory T cells Experiment Overall Design: Murine wild-type and mutant (delta) Foxp3 alleles were tagged with EGFP reporters. CD4 positive T cells expressing the respective tagged Foxp3 allele (Foxp3/EGFP and delta Foxp3/EGFP) were isolated from hemizygous male mice by cell sorting using FACS. CD4 positive cells that did not express the EGFP tag were a isolated. Total RNA was prepared and used in the array studies.
Project description:Localisation of CpG methylation in yeast expressing murine DNMTS Genomic DNA was purified from a control strain and a strain expressing murine DNMTs
Project description:Embryonically established osteoclast precursors expressing tdTomato reporter gene were analyzed by single cell RNA-sequencing. Data showed that hematopoietic stem cell independent yolk-sac erythromyeloid progenitors produced embryonic osteoclast precursors.