Project description:In order to determine that CUT&Tag is similar to known DUX ChIP-seq, we performed CUT&Tag with a mCherry-tagged DUX (with the mCherry antibody). Once confirmed, we pewrformed CUT&Tag for other DUX derivatives with their mCherry tag Then, we performed CUT&Tag for H3K9ac, which is known to globally increase in 2-cell-like cells, which occurs after DUX expression, and CUT&Tag for SMARCC1, a subunit of the SWI/SNF complex
Project description:We developed scNanoSeq-CUT&Tag, a streamlined method by adapting a modified CUT&Tag protocol to Oxford Nanopore sequencing platform for efficient chromatin modification profiling at single-cell resolution. We firstly tested the performance of scNanoSeq-CUT&Tag on six human cell lines: K562, 293T, GM12878, HG002, H9, HFF1 and adult mouse blood cells, it showed that scNanoSeq-CUT&Tag can accurately distinguish different cell types in vitro and in vivo. Moreover, scNanoSeq-CUT&Tag enables to effectively map the allele-specific epigenomic modifications in the human genome andallows to analyze co-occupancy of histone modifications. Taking advantage of long-read sequencing,scNanoSeq-CUT&Tag can sensitively detect epigenomic state of repetitive elements. In addition, by applying scNanoSeq-CUT&Tag to testicular cells of adult mouse B6D2F1, we demonstrated that scNanoSeq-CUT&Tag maps dynamic epigenetic state changes during mouse spermatogenesis. Finally, we exploited the epigenetic changes of human leukemia cell line K562 during DNA demethylation, it showed that NanoSeq-CUT&Tag can capture H3K27ac signals changes along DNA demethylation. Overall, we prove that scNanoSeq-CUT&Tag is a valuable tool for efficiently probing chromatin state changes within individual cells.
Project description:This study aimed to adapt CUT&Tag to Plasmodium falciparum samples as an efficient and sensitive alternative to classical ChIP-sequencing. We compare H3K9me3 and HP1 CUT&Tag with ChIP-seq datasets, showing successful establishment of CUT&Tag in P. falciparum. Next we aimed to scale down required input material for our CUT&Tag reactions and generated high-quality HP1 tracks with as little as 10.000 nuclei. To minimise potential sample loss we tested feasibility of utilising (frozen) saponin parasite isolates as input material instead of nuclei, which proved to be viable. Lastly, we deployed our new technique Dimerisation-induced Biotinylation-CUT&Tag (DiBioCUT&Tag) to catch transient interactions by biotinylation of strongly associated proteins such as histones. We tested this technique on HP1 and compared standart CUT&Tag with DiBioCUT&Tag. Furthermore, we explored interactions of the transcription factor BDP5, which we were previously unable to succesfully ChIP.
Project description:BRD4 inhibition suppressed M2 macrophage polarization. Particularly, we deeply investigated the underlying molecular mechanism of how BRD4 epigenetically regulate M2 genes expression. And finished the cut&tag experiment using BRD4, H3K27ac, IRF4 as antibody to find out the binding sites change in M2 macrophage treated with ARV825.
Project description:We recently introduced CUT&Tag, an epigenomic profiling strategy in which antibodies are bound to chromatin proteins in situ in permeabilized nuclei, and then used to tether the cut-and-paste transposase Tn5. Activation of the transposase simultaneously cleaves DNA and adds DNA sequencing adapters (“tagmentation”) for paired-end DNA sequencing. Here, we introduce a streamlined CUT&Tag protocol that suppresses exposure artifacts to ensure high-fidelity mapping of the antibody-targeted protein and improves signal-to-noise over current chromatin profiling methods. Streamlined CUT&Tag can be performed in a single PCR tube from cells to amplified libraries, providing low-cost high-resolution genome-wide chromatin maps. By simplifying library preparation, CUT&Tag requires less than a day at the bench from live cells to sequencing-ready barcoded libraries. Because of low background levels, barcoded and pooled CUT&Tag libraries can be sequenced for ~$25 per sample, enabling routine genome-wide profiling of chromatin proteins and modifications that requires no special skills or equipment.
Project description:Here we describe successful implementation of CUT&Tag for profiling protein-DNA interactions in zebrafish embryos. We optimized CUT&Tag protocol to generate high resolution maps of enrichment for the histone variant H2A.Z during zebrafish development. We were able to establish dynamics of H2A.Z genomic patterning from shield stage to 24hpf embryos. Our work demonstrates the power of combining CUT&Tag with the strengths of the zebrafish system to better understand the changing embryonic chromatin landscape and its roles in shaping development.
Project description:In this study, CUT&Tag-seq technology was employed to investigate MEF2A binding sites across the entire genome of chicken primary myoblasts. CUT&Tag was performed using CUT&Tag Assay Kit for Illumina Pro (TD904-1) from Vazyme. Antibody targeting MEF2A as well as IgG were used.The final DNA library on a HiSeq PE150 platform was subjected for the analyses. This study provides a wide landscape of MEF2A target genes from chicken primary myoblasts, which supports the active role of MEF2A in avian muscle development.
