Project description:Genome editing was conducted on a t(3;8) K562 model to investigate the effects of deleting different modules or CTCF binding sites within the MYC super-enhancer. To check mutations after targeting with CRISPR-Cas9 we performed amplicon sequencing using the Illumina PCR-based custom amplicon sequencing method using the TruSeq Custom Amplicon index kit (Illumina). The first PCR was performed using Q5 polymerase (NEB), the second nested PCR with KAPA HiFi HotStart Ready mix (Roche). Samples were sequenced paired-end (2x 250bp) on a MiSeq (Illumina).
Project description:Chromatin immunoprecipitation analysis of CENH3 in the Arabidopsis thaliana accessions Col-0, Ler-0, Cvi-0 and Tanz-1 was performed in order to align reads to PacBio HiFi genome assemblies which contain complete centromere repeat arrays.
Project description:Understanding transcriptional regulatory networks (TRNs) in microglia is key to uncovering mechanisms driving central nervous system (CNS) disorders. Human iPSC-derived models offer a tractable system for studying microglia, yet variability between lines has limited reproducibility. Here, we use the standardized KOLF2.1J iTF line to rapidly generate microglia-like cells (iTF-Microglia) and profile TRNs under homeostatic and inflammatory conditions. iTF-Microglia closely resemble primary brain microglia at both transcriptomic and epigenomic levels. Integrative analyses reveal microglia-enriched candidate cis-regulatory elements (cCREs) and dynamic enhancer remodeling upon differentiation and LPS+IFNG stimulation, involving key transcription factors (TFs) including NF-kB, IRF, and STAT families. TRNs active in iTF-Microglia are enriched for genetic variants linked to Alzheimer’s disease and other CNS disorders. These findings establish KOLF2.1J iTF-Microglia as a reproducible and genetically tractable platform for studying human microglial gene regulation and provide mechanistic insight into how TRN remodeling may contribute to CNS disease risk.
Project description:Understanding transcriptional regulatory networks (TRNs) in microglia is key to uncovering mechanisms driving central nervous system (CNS) disorders. Human iPSC-derived models offer a tractable system for studying microglia, yet variability between lines has limited reproducibility. Here, we use the standardized KOLF2.1J iTF line to rapidly generate microglia-like cells (iTF-Microglia) and profile TRNs under homeostatic and inflammatory conditions. iTF-Microglia closely resemble primary brain microglia at both transcriptomic and epigenomic levels. Integrative analyses reveal microglia-enriched candidate cis-regulatory elements (cCREs) and dynamic enhancer remodeling upon differentiation and LPS+IFNG stimulation, involving key transcription factors (TFs) including NF-kB, IRF, and STAT families. TRNs active in iTF-Microglia are enriched for genetic variants linked to Alzheimer’s disease and other CNS disorders. These findings establish KOLF2.1J iTF-Microglia as a reproducible and genetically tractable platform for studying human microglial gene regulation and provide mechanistic insight into how TRN remodeling may contribute to CNS disease risk.
Project description:Understanding transcriptional regulatory networks (TRNs) in microglia is key to uncovering mechanisms driving central nervous system (CNS) disorders. Human iPSC-derived models offer a tractable system for studying microglia, yet variability between lines has limited reproducibility. Here, we use the standardized KOLF2.1J iTF line to rapidly generate microglia-like cells (iTF-Microglia) and profile TRNs under homeostatic and inflammatory conditions. iTF-Microglia closely resemble primary brain microglia at both transcriptomic and epigenomic levels. Integrative analyses reveal microglia-enriched candidate cis-regulatory elements (cCREs) and dynamic enhancer remodeling upon differentiation and LPS+IFNG stimulation, involving key transcription factors (TFs) including NF-kB, IRF, and STAT families. TRNs active in iTF-Microglia are enriched for genetic variants linked to Alzheimer’s disease and other CNS disorders. These findings establish KOLF2.1J iTF-Microglia as a reproducible and genetically tractable platform for studying human microglial gene regulation and provide mechanistic insight into how TRN remodeling may contribute to CNS disease risk.
Project description:Understanding transcriptional regulatory networks (TRNs) in microglia is key to uncovering mechanisms driving central nervous system (CNS) disorders. Human iPSC-derived models offer a tractable system for studying microglia, yet variability between lines has limited reproducibility. Here, we use the standardized KOLF2.1J iTF line to rapidly generate microglia-like cells (iTF-Microglia) and profile TRNs under homeostatic and inflammatory conditions. iTF-Microglia closely resemble primary brain microglia at both transcriptomic and epigenomic levels. Integrative analyses reveal microglia-enriched candidate cis-regulatory elements (cCREs) and dynamic enhancer remodeling upon differentiation and LPS+IFNG stimulation, involving key transcription factors (TFs) including NF-kB, IRF, and STAT families. TRNs active in iTF-Microglia are enriched for genetic variants linked to Alzheimer’s disease and other CNS disorders. These findings establish KOLF2.1J iTF-Microglia as a reproducible and genetically tractable platform for studying human microglial gene regulation and provide mechanistic insight into how TRN remodeling may contribute to CNS disease risk.