Project description:RNA sequencing of FACS sorted microglia isolated from brain tissue of WT, Hexb-tdTomato, Hexb-CreERT2 and Hexb-KO mice was performed to test for potential differences in gene expression caused by the knock-in of tdTomato and CreERT2 into the Hexb locus.
Project description:RNA sequencing of FACS sorted microglia isolated from brain tissue of WT, Hexb-tdTomato, Hexb-CreERT2 and Hexb-KO mice was performed to test for potential differences in gene expression caused by the knock-in of tdTomato and CreERT2 into the Hexb locus.
Project description:RNA sequencing of FACS sorted microglia isolated from brain tissue of WT, Hexb-tdTomato, Hexb-CreERT2 and Hexb-KO mice was performed to test for potential differences in gene expression caused by the knock-in of tdTomato and CreERT2 into the Hexb locus.
Project description:All current smooth muscle cell (SMC) restricted Cre mice recombine floxed alleles in vascular and visceral SMCs. We generated a tamoxifen-inducible CreERT2 mouse, Itga8-CreERT2, and compared its activity to the widely used Myh11-CreERT2 mouse. Both CreERT2 mice showed similar activity in vascular SMCs; however, Itga8-CreERT2 displayed low activity in visceral SMC-containing tissues (e.g., intestine). Myh11-CreERT2 (but not Itga8-CreERT2) mice exhibited high levels of CreERT2 protein, tamoxifen-independent activity, and an altered transcriptome. Whereas Myh11-CreERT2-mediated knockout of Srf resulted in a lethal intestinal phenotype, loss of Srf with Itga8-CreERT2 (SrfItga8) revealed viable mice with attenuated vascular SMC contractile gene expression, but no evidence of intestinal pathology. Male and female SrfItga8 mice presented with vascular contractile incompetence; however, only male SrfItga8 mice showed systemic changes in blood pressure. These results establish the Itga8-CreERT2 mouse as an alternative to existing SMC Cre strains where gene loss may result in visceral myopathies that obfuscate accurate phenotyping in vascular SMCs.
Project description:We used microfluidic single cell RNA-seq on adult isolated CC10-CreERT2 (negative) integrin beta4(pos) cells lung epithelial cells in order to determine the transcriptional profile of this putative progenitor population. CC10-CreERT2 / tdTomato (negative) integrin beta4(pos) cells were isolated by FACS, as were Krt5-CreERT2 / tdTomato (positive) cells. These cells were pooled and loaded onto the Fluidigm C1 device.
Project description:Fluorescent cerebella were subsequently dissected at E15.5, E17.5, P0 (day of birth), P7 (postnatal day 7), P14 and P30; and tdTomato+ CGNPs were purified using fluorescence activated cell sorting (FACS). From these samples, we generated transcriptomes using RNA-seq
Project description:To investigate the mechanisms that underlie astrocyte dedifferentiation, we performed single cell RNA sequencing analysis of tdTomato+ astrocyte derived cells approx 1 year after stab-wound injury. 2-3 month old GFAP-CreERT2;p53flox/flox;LSL-tdTomato (p53Gfap-icKO) mice were subjected to intracortical endoxifen injection to enable deletion of p53 gene and simultaneous expression of tdTomato for fate-mapping specifically in astrocytes (GFAP+) in the context of a stab-wound injury. GFAP-CreERT2;p53wt/wt;LSL-tdTomato (p53Gfapwt/wt) mice were used as control whereby p53 remains fate-mapped tdTomato+ astrocytes remain p53 wildtype after endoxifen injection. Approx 10-12 months after injury, mice were sacrificed and the peri-wound region was dissected. Mice of the same genotypes were pooled (3 p53Gfap-icKO and 2 p53Gfapwt/wt) and digested using the Miltenyi Adult brain dissociation kit. Single cell suspension was FACS sorted for tdTomato+ cells into 384 well plates and then Smartseq3 libraries were generated.