Project description:To reveal the role of NCOA7 in cellular senescence, we performed the CUT&TAG assay using the H3K27ac antibody to map acetylation in granulosa cells from control and POI patients. We conducted the DNA sequencing of libraries from CUT&TAG assay using the H3K27ac antibody in human primary granulosa cells.
Project description:To reveal the role of MCM8 in suppressing R-loop accumulation, we performed the CUT&TAG assay using the S9.6 antibody to map genome-wide R-loops in Mcm8 wildtype MEFs and Mcm8 knockout MEFs. We also conducted the CUT&TAG assay to detect genome-wide R-loops in Ddx5 downregulated MEFs by adenovirus infection and in control MEFs. To investigate the underlying molecular mechanism of MCM8 suppressing R-loops, we conducted the DNA sequencing of libraries from CUT&TAG assay using the antibody against FLAG in HEK293 cells transfected with FLAG-MCM8 plasmid and using the S9.6 antibody in HEK293 cells. Besides, an IgG control and control of RNH1 overexpression were included.
Project description:To gain mechanistic insight into how Epigenetic factors reprogramming metabolism in response to DON treatment. we performed CUT&Tag sequencing in murine PDAC cells. sgPaxip1 cells were treated in Cont group or DON group , and harvested after 72 hours. CUT&Tag assay was performed following the manual of hyperactive pG-Tn5/pA-Tn5 transposase for CUT&Tag kit (TD901, Vazyme). DNA library were prepared according to manufacturer’s instructions of Trueprep index kit v2 (TD202, Vazyme).
Project description:To gain mechanistic insight into how Epigenetic factors reprogramming metabolism in response to glutamine starvation. we performed CUT&Tag sequencing in murine PDAC cells. KPC1199 cells were treated in Cont group or Low Gln group , and harvested after 72 hours. CUT&Tag assay was performed following the manual of hyperactive pG-Tn5/pA-Tn5 transposase for CUT&Tag kit (TD901, Vazyme). DNA library were prepared according to manufacturer’s instructions of Trueprep index kit v2 (TD202, Vazyme).
Project description:To reveal the role of MCM8 in suppressing R-loop accumulation, we performed the CUT&TAG assay using the S9.6 antibody to map genome-wide R-loops in MEFs derived from Mcm8 knockout and wildtype embryos. To investigate the underlying molecular mechanism of MCM8 suppressing R-loops, we conducted the DNA sequencing of libraries from CUT&TAG assay using the antibody against FLAG in HEK293 cells transfected with FLAG-MCM8 plasmid and using the S9.6 antibody in HEK293 cells.
Project description:We performed the cleavage under targets and tagmentation (Cut & tag) assay followed by sequencing enriched DNA fragments to reveal the direct downstream targets of Pbx1. Firstly, we overexpressed Pbx1b with Pbx1b-IRES-GFP retrovirus in murine peripheral B cells to ensure the yields of DNA fragments. CUT & tag libraries were generated following instructions of the manufacturer’s protocol (Vazyme; cat TD901-01) and the Pbx1 antibody (CST; cat 4342) was used for signal enrichment.
Project description:CUT&Tag was performed to identify the effect of DIS3 depletion on DNA:RNA hybrids genome-wide. DIS3-AID HCT116 cells were treated with either Auxin (for rapid depletion of DIS3) or DMSO (control). CUT&Tag was performed targeting DNA:RNA hybrids using the CUT&Tag-IT® R-loop Assay Kit (Active Motive) with the S9.6 antibody.
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.
