Journal: Genome Biology

Article Title: Identification of X-chromosomal genes that drive sex differences in embryonic stem cells through a hierarchical CRISPR screening approach

doi: 10.1186/s13059-021-02321-2

Figure Lengend Snippet: Over-expression of Klhl13 and Dusp9 in male mESCs leads to an enhanced pluripotency state and slower differentiation kinetics. a Schematic representation of the dCas9-SunTag system used for gene activation. b–e To over-express Dusp9 (yellow) and Klhl13 (blue), male E14 mESCs, stably expressing the doxycycline-inducible SunTag system, were either transduced with one of two different sgRNAs targeting the respective promoter regions or with non-targeting control (NT) sgRNAs and were treated for 3 days with 1 μg/ml doxycycline as indicated. Protein levels of Dusp9 (left) and Klhl13 (right) were quantified via immunoblotting ( b ), expression levels of MAPK target genes Spry4 and Egr1 ( c ) and of naive pluripotency factors Nanog and Prdm14 ( e ) were assessed by qPCR and phosphorylation of Mek and Erk was quantified by immunoblotting ( d ). The immunoblot signals were normalized to Tubulin ( b ) or to total Mek/Erk ( d ) and to the mean of two doxycycline-treated non-targeting control sgRNAs. qPCR measurements were normalized to two housekeeping genes and to the respective untreated control (−Dox). Dots and triangles depict individual measurements of the two different sgRNAs, and thick bars show the mean of three biological replicates. f Dusp9- and Klhl13 over-expressing mESCs were treated with 1 μg/ml doxycycline 24 h before differentiation via LIF withdrawal for 4 days, and expression levels of pluripotency factors were measured by qPCR at different time points as indicated. Mean and standard deviation across 3 biological replicates is shown. g Global CpG methylation levels in cell lines over-expressing Dusp9 and Klhl13 via doxycycline treatment for 3 passages were assessed via pyrosequencing-based luminometric DNA methylation assay (LUMA). * p < 0.05 in a two-tailed paired Student’s t test comparing the Dusp9/Klhl13 over-expressing samples and the non-targeting controls (mean of sgRNA1 and sgRNA2)

Article Snippet: Mean and standard deviation across 3 biological replicates is shown. g Global CpG methylation levels in cell lines over-expressing Dusp9 and Klhl13 via doxycycline treatment for 3 passages were assessed via pyrosequencing-based luminometric DNA methylation assay (LUMA).

Techniques: Over Expression, Activation Assay, Stable Transfection, Expressing, Transduction, Control, Western Blot, Phospho-proteomics, Standard Deviation, CpG Methylation Assay, DNA Methylation Assay, Two Tailed Test

Journal: Cancers

Article Title: Impact of Physical Activity on DNA Methylation Signatures in Breast Cancer Patients: A Systematic Review with Bioinformatic Analysis

doi: 10.3390/cancers16173067

Figure Lengend Snippet: Effects of PA on DNA methylation patterns in BC populations. Table summarising the study design and results of interest from all selected articles that evaluated the effects of PA on DNA methylation signatures in BC patients. DNA methylation results are presented using the gene codes of the genes associated and/or affected by the PA-induced methylation changes reported by the original authors.

Article Snippet: The authors also analysed global DNA methylation in white blood cell DNA [ , ] using the luminometric methylation assay (LUMA), a quantitative measurement of genome-wide DNA methylation, as described by Bjornsson et al. [ ], and LINE-1, where four CpG sites in the promoter region of LINE-1 were assessed using a validated pyrosequencing-based methylation assay [ ].

Techniques: DNA Methylation Assay, Methylation, Control

Journal: Environmental Epigenetics

Article Title: Epigenetic processes involved in response to pesticide exposure in human populations: a systematic review and meta-analysis

doi: 10.1093/eep/dvae005

Figure Lengend Snippet: Articles assessing global DNA methylation in response to pesticide exposure

Article Snippet: Itoh et al ., [ ] , Cross-sectional , DNA methylation in peripheral leukocytes , LUMA (pyrosequencing) , 403 Japanese women , High-resolution mass spectrometer with selected ion monitoring coupled to a gas chromatograph, based on isotope dilution mass spectrometry , β-HCH, pp' -DDE, cis -heptachlor epoxide, trans -nonachlor ( trans -nonachlordane), pp' -DDD (dichlorodiphenyldichloroethane, pp' -DDT dichlorodiphenyltrichloroethane , Global hypomethylation.

Techniques: DNA Methylation Assay, Pesticides, Mass Spectrometry, Isotope Dilution, Liquid Chromatography, Gas Chromatography, Clinical Proteomics, Gas Chromatography-Mass Spectrometry, Methylation, Chromatography