Tag: K562

Pan-hematopoietic Tox4 deletion reduces number of multipotential progenitors and impairs T cell development To understand the role of TOX4 in development, we generated Tox4 conditional knockout mice by the CRISPR-Cas9 methodology, and two loxP sites in the same orientation were inserted upstream and downstream of exons 4–6, respectively (Supplementary Fig. 1a). Considering…

In a recent study published in Nature Metabolism, researchers attempted to identify new genes and molecular pathways that might supply energy when the availability of glucose or other nutrients is limited. Study: Salvage of ribose from uridine or RNA supports glycolysis in nutrient-limited conditions. Image Credit: Irina Anosova/Shutterstock.com Background In…

Modification of HDRTs with interstrand crosslinks increases HR during gene editing. (a) Top panel: Cas9 RNPs introduce a double strand DNA break (DSB) at a targeted region in the genome, which can be repaired by error prone end joining (EJ) processes that rejoin the ends of the break, or homology-directed…

Cells and cell culture KOPN30, BV173, and K562 are BCR/ABL1-positive leukemia cell lines. All leukemia cell lines, as well as Ba/F3 cells, were maintained in RPMI-1640 medium supplemented with 15% fetal bovine serum (FBS) and penicillin–streptomycin (100 U/mL) at 37 °C in an atmosphere containing 5% CO2. KOPN30 cells were obtained…

CLIP enables high-efficiency knock-in into an essential locus To create CLIP, we leverage integrase-deficient lentiviruses (IDLV) to deliver an HDR donor template as a viral RNA genome to cells25,26. IDLV consists of lentiviral components but with a D64V mutation in the integrase, maintaining this enzyme’s ability to package and nuclear…

Human CD34+ cell isolation Human fetal liver (FL) samples were obtained from Advanced Biosciences Resources (Alameda, CA) with proper consent. FL samples were cut in small fragments, treated for 25 min at 37 °C with Collagenase D (100 ng/mL; Roche). The cell suspension was prepared, and the human CD34+ cells were separated by…

The present protocol shows how to differentiate CD3−/CD45+CD56+ cells with mild cytotoxicity from human expanded potential stem cells (hEPSCs) under both 3D and 2D culture conditions. This allows for routine phenotypical validation without the destruction of the complex microenvironment. Speaking of significance, generating NK cells from an expandable source is…

Multiple H2BNTac sites occupy the same genomic regions H2BNTac sites are similarly regulated by CBP/p30018 (Supplementary Fig. 1a), yet the reported genome occupancy patterns of H2BNTac sites are dissimilar from each other22,23 (Supplementary Note 1). To resolve this conundrum, we systematically compared H3K27ac and H2BNTac genomic occupancy and regulation by…

FG spectrum in pediatric AML patients A combinatorial approach, including conventional and next-generation sequencing (NGS)-based assays, was employed to profile the FG landscape (see Methods). FG analysis was feasible for 138 patients (94% of the entire cohort, Supplementary Data 1) from whom suitable testing materials were available. The most common FGs…

Abstract CRISPR-based gene activation (CRISPRa) is a promising therapeutic approach for gene therapy, upregulating gene expression by targeting promoters or enhancers in a tissue/cell-type specific manner. Here, we describe an experimental framework that combines highly multiplexed perturbations with single-cell RNA sequencing (sc-RNA-seq) to identify cell-type-specific, CRISPRa-responsive cis-regulatory elements and the…

ANKLE1 is the causal gene for breast and ovarian cancer risk in the chr19p13.1 region Expression quantitative trait loci (eQTL) data have revolutionized how geneticists identify candidate causal genes from genome-wide association study (GWAS) loci. We integrated the most recent meta-analysis of breast cancer GWAS10 and Genotype-Tissue Expression (GTEx) project…

Sponsored Content by TecanJan 25 2023 Making Double-stranded DNA Quantification and Normalization easy and fast using Promega QuantiFluor® dyes with the Fluent® Laboratory Automation Solution from Tecan. Introduction The field of genomics demands double-stranded (ds) DNA quantification and normalization to a high standard of reliability. While absorbance techniques have been…

Abstract Nuclear noncoding RNAs (ncRNAs) are key regulators of gene expression and chromatin organization. The progress in studying nuclear ncRNAs depends on the ability to identify the genome-wide spectrum of contacts of ncRNAs with chromatin. To address this question, a panel of RNA–DNA proximity ligation techniques has been developed. However,…

Some genome editing systems are highly conspicuous. They introduce double-strand breaks to DNA that attract the attention of cellular mechanisms such as nonhomologous end joining and homology-directed repair. If a genome editing system is so brash as to attempt a sizable insertion of new DNA, homology-directed repair must ensure that…

CANCER & SINGLE-CELL ANALYSIS The heterogeneity and dynamism of cancer present formidable challenges to understanding and treating the disease. As discussed in the last section, tumor evolution often leads to considerable genetic variation across clones. But the complexity of tumors does not stop at the level of DNA. Intratumoral phenotypic…