Project description:Phenol Toluol extraction (PTex) carried out two steps organic-phase separation: first pH neutral phenol–Toluol extraction separated RNA and protein from DNA: RNA and proteins to the top aqueous phase but DNA and membranes in the interface. Aqueous phase then went through two rounds acid-phenol extraction, resulting RNA in the top aqueous phase, proteins bottom organic phase and RNA-protein complexes in the interface. After carefully isolated the interface, ethanol precipitation was carried on for purifying RNA-protein adducts (Urdaneta et al, 2019; Smith et al, 2020)
2022-05-31 | PXD023414 | Pride
Project description:Comparison of DNA extraction methods for long-read mNGS testing
| PRJNA824617 | ENA
Project description:Comparison of DNA extraction methods for long reading mNGS testing
Project description:While DNA methylation is an important gene regulatory mechanism in mammals (Razin and Riggs 1980; Moore, Le, and Fan 2013), its function in arthropods remains poorly understood. Studies in eusocial insects have argued for its role in caste development by regulating gene expression and splicing (Elango et al. 2009; Lyko et al. 2010; Bonasio et al. 2012; Flores et al. 2012; Foret et al. 2012; Li-Byarlay et al. 2013; Marshall, Lonsdale, and Mallon 2019; Shi et al. 2013)(Alvarado et al. 2015; Kucharski et al. 2008). However, such findings are not always consistent across studies, and have therefore remained controversial (Arsenault, Hunt, and Rehan 2018; Cardoso-Junior et al. 2021; Harris et al. 2019; Herb et al. 2012; Libbrecht et al. 2016; Oldroyd and Yagound 2021b; Patalano et al. 2015). Here we use CRISPR/Cas9 to mutate the maintenance DNA methyltransferase DNMT1 in the clonal raider ant, Ooceraea biroi. Mutants have greatly reduced DNA methylation but no obvious developmental phenotypes, demonstrating that, unlike mammals (Brown and Robertson 2007; En Li, Bestor, and Jaenisch 1992; Jackson-Grusby et al. 2001; Panning and Jaenisch 1996), ants can undergo normal development without DNMT1 or DNA methylation. Additionally, we find no evidence of DNA methylation regulating caste development. However, mutants are sterile, while in wildtypes, DNMT1 is localized to the ovaries and maternally provisioned into nascent oocytes. This supports the idea that DNMT1 plays a crucial but unknown role in the insect germline (Amukamara et al. 2020; Arsala et al. 2021; Bewick et al. 2019; Schulz et al. 2018; Ventós-Alfonso et al. 2020; Washington et al. 2020).
Project description:RNAseq of CD14 cells from 60 female RA patients activated with LPS for 2 hours. Patients were collected crossectionally from the Rheumatology clinic during 2019-2020. Age md 64, range 23-76 years, Disease duration md 10, range 1-45 years.
Project description:RNAseq of CD4 cells from 60 female RA patients activated with aCD3 for 2 hours. Patients were collected crossectionally from the Rheumatology clinic during 2019-2020. Age md 64, range 23-76 years, Disease duration md 10, range 1-45 years.
Project description:We report a method for specific capture of an arbitrary subset of genomic targets for single molecule bisulfite sequencing, and for digital quantitation of DNA methylation at a single nucleotide resolution. We used targeted bisulfite sequencing to characterize the changes of DNA methylation during the de-differentiation of human fibroblasts into hybrid stem cells, and into induced pluripotent stem cells. We compared the methylation level of approximately 66,000 CpG sites within 2020 CpG islands on chromosome 12, chromosome 20, and 34 selected regions. A total of 288 differentially methylated regions were identified between fibroblasts and pluripotent cells. Methylation cluster analysis revealed distinct methylation patterns between fibroblasts and pluripotent cells. Furthermore iPS cells are globally more methylated than human embryonic stem cells, which could be due to the reprogramming process. This targeted bisulfite sequencing method is particularly useful for efficient and large-scale analysis of DNA methylation in organisms with large genomes. Experiment Overall Design: Comparison of DNA methylation on 2020 CpG islands and 34 other selected regions among eleven human ES, iPS and fibroblast lines.