Project description:In this study, we present a multiplexed version of Deterministic Barcoding in Tissue (xDBiT) to acquire spatially resolved transcriptomes of nine tissue sections in parallel. New microfluidic chips were developed to spatially encode mRNAs over a total tissue area of 1.17 cm2 with spots of 50 µm×50 µm. Optimization of the biochemical protocol increased read and gene counts per spot by one order of magnitude compared with previous reports. Furthermore, the introduction of alignment markers allows seamless registration of images and spatial transcriptomic spot coordinates. Together with technological advances, we provide an open-source computational pipeline to transform raw sequencing data from xDBiT experiments into the AnnData format. The functionality of xDBiT was demonstrated by the acquisition of 16 spatially resolved transcriptomic datasets from five different murine organs, including cerebellum, liver, kidney, spleen, and heart. Factor analysis and deconvolution of xDBiT spatial transcriptomes allowed for in-depth characterization of the murine kidney.
Project description:Multiplexed transcriptome-based profiling of drug combinations using deterministic barcoding in picolitre-sized droplets [small_scale_drug_screen]
Project description:Multiplexed transcriptome-based profiling of drug combinations using deterministic barcoding in picolitre-sized droplets [large_scale_drug_screen]
Project description:It is elusive whether clonal selection of tumor cells in response to ionizing radiation (IR) is a deterministic or stochastic process. With high resolution clonal barcoding and tracking of over 400.000 HNSCC patient-derived tumor cells the clonal dynamics of tumor cells in response to IR was analysed. Fractionated IR induced a strong selective pressure for clonal reduction. This significantly exceeded uniform clonal survival probabilities indicative for a strong clone-to clone difference within tumor cells. Survival to IR is driven by a deterministic clonal selection of a smaller population which commonly survives radiation, while increased clonogenic capacity is a result of clonal competition of cells which have been selected stochastically. The ratio of these parameters is amenable to radiation sensitivity which correlates to prognostic biomarkers of HNSCC. Evidence for the existence of a rare subpopulation with an intrinsically radiation resistant phenotype was found at a frequency of 0.6-3.3%. With cellular barcoding we introduce a novel functional heterogeneity associated qualitative readout for evaluating the contribution of stochastic and deterministic clonal selection processes in response to IR.
Project description:Multiplexed transcriptome-based profiling of drug combinations using deterministic barcoding in picolitre-sized droplets [dataset 3]
Project description:It is elusive whether clonal selection of tumor cells in response to ionizing radiation (IR) is a deterministic or stochastic process. With high resolution clonal barcoding and tracking of over 400.000 HNSCC patient-derived tumor cells the clonal dynamics of tumor cells in response to IR was analysed. Fractionated IR induced a strong selective pressure for clonal reduction. This significantly exceeded uniform clonal survival probabilities indicative for a strong clone-to clone difference. within tumor cells. Survival to IR is driven by a deterministic clonal selection of a smaller population which commonly survives radiation, while increased clonogenic capacity is a result of clonal competition of cells which have been selected stochastically. The ratio of these parameters is amenable to radiation sensitivity which correlates to prognostic biomarkers of HNSCC. Evidence for the existence of a rare subpopulation with an intrinsically radiation resistant phenotype was found at a frequency of 0.6-3.3%. With cellular barcoding we introduce a novel functional heterogeneity associated qualitative readout for evaluating the contribution of stochastic and deterministic clonal selection processes in response to IR. To analyze transcriptomic changes of HNSCC cell lines after fractionated Photon IR (5x4Gy), RNAseq analysis was performed on irradiated cells in comparison to untreated control cells (EBI submission E-MTAB-9693)
Project description:We present spatially resolved high-spatial-resolution genome-wide co-mapping of epigenome and transcriptome by simultaneously profiling of chromatin accessibility and gene expression (spatial-ATAC-RNA-seq), as well as histone modification and gene expression (spatial-CUT&Tag-RNA-seq) on the same tissue section at cellular level by combining the microfluidic deterministic barcoding strategy in DBiT-seq and the chemistry used in ATAC-seq/CUT&Tag.
Project description:We present spatially resolved chromatin accessibility profiling of tissue sections via next-generation sequencing by combining in situ ATAC-seq chemistry and microfluidic deterministic barcoding. Mouse and human tissues were profiled by spatial-ATAC-seq.