Genome-wide expression profiles from mouse Phf13 wildtype and knockout testes
ABSTRACT: We report expression profiles for mouse testes carrying a Phf13 knockout in comparison to wildtype testes. Upregulation of X- and Y-linked genes is evident after loss of PHF13. Overall design: RNA-Seq of 3 samples each from wildtype and Phf13 knockout mice.
Project description:CG3875 is a young duplicate gene of kep1 family originated recently in Drosophila melanogaster (D. melanogaster) species complex (including D. melanogaster, D. simulans, D. mauritiana and D. sechellia) after it split from D. yakuba. It encodes an RNA-binding protein containing a single maxi-KH domain. Characterization of loss-of-function phenotypes of CG3875 mutants generated by gene targeting demonstrated that CG3875 null males display a seriously reduced fertility compared with wildtype males and most of the null males are completely sterile. Further cytological identification of CG3875 null males suggested that CG3875 plays an important role in spermiogenesis processes including sperm individualization and sperm coiling. In addition, CG3875 is also essential for the formation of outer dynein arm of sperm axoneme. In order to identify the molecular mechanism responsible for the involvement of CG3875 in spermiogenesis and structural integrity of sperm axoneme, we performed microarray analysis to identify transcripts whose levels are altered in the testes of CG3875 null males. Overall design: We compared gene expression profiles between testes of 0-2 day CG3875 null mutants and wildtype flies, respectively. For the sake of an identical genetic background between knockout flies and wildtype flies, the wildtype stock used here is wildtype recombinant line created during gene targeting. Seminal vesicles were removed from the testes during dissection. Two independent RNA extractions and hybridizations were conducted for each sample.
Project description:In situ synthesized oligo arrays, U74Av2, from Affymetrix were used to measure differential gene expression in RNA samples generated from the liver (GSE864) and spleen (GSE865) of Nrf2 knockout and wildtype mice at 5 months of age. This SuperSeries is composed of the following subset Series:; GSE864: Nrf2 Knockout vs. Wildtype mice, liver sample; GSE865: Nrf2 Knockout vs. Wildtype mice, spleen sample
Project description:Ikbkap/Elp1 regulates genes involved in spermatogenesis. Ikbkap deficiency causes male infertility by disrupting meiotic progression. In this dataset, we include the expression data for control and Ikbkap-knockout mouse testis. We analyzed the gene expression in control and Ikbkap-knockout testes using the the Affymetrix MoGene-1_0-st-v1 platform. Three replicates per genotype.
Project description:We examined the impact of early-life exposure to methylmercury (MeHg) on Caenorhabditis elegans (C. elegans) pdr-1 mutants, addressing gene-environment interactions. We tested the hypothesis that early-life exposure to MeHg and knockout (KO) of pdr-1 (mammalian: parkin/PARK2) exacerbates MeHg toxicity and damage to the dopaminergic (DAergic) system. pdr-1KO worms showed increased lethality and decreased lifespan following MeHg exposure. Mercury (Hg) content, measured with inductively coupled plasma-mass spectrometry was increased in pdr-1KO worms compared to wildtype (N2) controls. 2'7' dichlorodihydrofluorescein diacetate assay revealed a significant increase in reactive oxygen species in both strains following MeHg exposure; however, while N2 worms showed an increase in skn-1 transcript levels following MeHg exposure, there was no difference in skn-1 induction in pdr-1KO worms. Dopamine-dependent behavioral analysis revealed an effect of MeHg on N2 wildtype worms, but no effect on pdr-1KO worms. Taken together, these results suggest that pdr-1KO worms are more sensitive to MeHg than wildtype worms, but MeHg does not exacerbate behavioral changes related to the absence of pdr-1.
Project description:A fluorescence marker mOrange was inserted to the popular pLentiCrispr-V2 to create pLentiCrispr-V2-mOrange (V2mO) that contained both a puromycin selection and a fluorescent marker, making viral production and target transduction visible. Lentiviruses packaged with this plasmid and appropriate guide RNAs (gRNAs) successfully knocked out the genes RhoA, Gli1, and Gal3 in human gastric cancer cell lines. Cas9-gRNA editing efficiency could be estimated directly from Sanger electropherograms of short polymerase chain reaction products around the gRNA regions in Cas9-gRNA transduced cells. Single cloning of transduced target cell pools must be performed to establish stable knockout clones. Rescue of wildtype (RhoA and Gal3) and mutant (RhoA.Y42C) genes into knockout cells was successful only when cDNAs, where gRNAs bind, were modified by three nucleotides while the amino acid sequences remained unchanged. Stringent on-target CRISPR/Cas9 editing was observed in Gal3 gene, but not in RhoA gene since RhoA.Y42C already presented a nucleotide change in gRNA5 binding site. In summary, our improved strategy added these advantages: adding visual marker to the popular lentiviral system, monitoring lentiviral production and transduction efficiencies, cell-sorting Cas9+ cells in target cells by fluorescence-activated cell sorting, direct estimation of gene editing efficiency of target cell pools by short PCR electropherograms around gRNA binding sites, and successful rescue of wildtype and mutant genes in knockout cells, overcoming Cas9 editing by modifying cDNAs.
Project description:Expression changes in testes of 5 week old mice after knockout of Phf13 were analyzed using Affymetrix mouse genome 430 2.0 expression microarrays. Transcripts on the X- and Y-chromosome were significantly upregulated. Overall design: RNA was isolated from the testes of 5 week old PHF13-knockout and wild type mice and analyzed using Affymetrix expression microarrays.
Project description:Expression changes in testes of 15-20 week old mice after knockout of Phf13 were analyzed using Affymetrix mouse genome 430 2.0 expression microarrays. Transcripts on the X- and Y-chromosome were significantly upregulated. Overall design: RNA was isolated from the testes of 15-20 week old PHF13-knockout and wild type mice and analyzed using Affymetrix expression microarrays.
Project description:Foxl2 is a forkhead transcription factor expressed only in the female, but not in the male gonad. We have created mice homozygous mutant for the Foxl2 gene (KO) as well as mice carrying a conditional mutant Foxl2 allele (floxed). The expression profiles of conventional Foxl2 knockout and wildtype ovaries were compared at P3, using the Affy Mouse Genome 430 2.0 Array. Adult wildtype and conditional mutant (Foxl2 floxed x RosaCre-EBD treated with tamoxifen) ovaries were compared to adult wildtype testes using the Affymetrix Mouse Gene 1.0 ST Array. Both experiments (KO/WT P3 and Mutant/WT/Testis Adult were also compared to each other.) Overall design: Ovaries from newborn (postnatal day 3) and adult (8 week old) mice were excised from the animal and RNA was prepared from whole ovaries (Adult testes were excised and RNA was isolated from 1/4 testis).