Project description:Mouse Wdr74-knockout embryos were generated using the CRISPR-Cas9 system. Embryos from the 4-cell stage to the morula stage were collected and treated with acidic Tyrode's solution to remove the zona pellucida. We used 50 embryos per sample at each developmental stage and identified the proteome of mouse Wdr74-knockout embryos by label-free quantification.
Project description:WT immortorized mouse macrophages and CISPR CAS 9 mediated Myristoylated, Alanine-rich C-kinase Substrate knockout ΔMARCKS were treated with LPS for 1h, 6h, 24h. No treatment both WT and ΔMARCKS were use as a control experiment. The label free quantification were prefromed using
Project description:Methods that enable absolute quantification of N6-methyladenosine (m6A) RNA modification have emerged as powerful tools in the field of epitranscriptomics. We previously reported GLORI, a chemical-assisted approach firstly achieved quantitatively transcriptome-wide m6A measurement at single-base resolution. Despite its advantages, GLORI suffers from lengthy reaction time and severe RNA degradation. Here, we present two updated GLORI approaches: GLORI 2.0 is an ultra-fast and mild version that preserves RNA integrity and enhances sensitivity for both transcriptome-wide and locus-specific m6A detection; GLORI 3.0 further utilizes a novel reverse transcription-silent carrier RNA to achieve high-quality m6A quantification from ~ 1,000 cells. Using limited RNA input extracted from single mouse dorsal hippocampus, we measure m6A methylome in the synaptic and cytoplasmic fractions and reveal a high modification level in synapse-related gene sets. We envision that the updated GLORI methods will greatly expand the applicability of absolute quantification of m6A in biology.