Cellular Multiverse In A Nutshell: Advanced Single Cell Multi-Omics Solutions 23.11.20225’ Single cell sequencing Cellular Multiverse In A Nutshell: Advanced Single Cell Multi-Omics SolutionsRecently, single cell transcriptome sequencing technology has rapidly developed to provide effective methods for researchers to study the heterogeneity of cells; to understand molecular mechanisms and developmental processes; and to investigate different disease states at a single cell resolution. Here we present novel multi-omics methods that combine transcriptomic data with additional information on diverse analytes to expand insights into the molecular mechanisms of cellular functions. Figure 1. Summary of the unique solutions offered by Singleron to advance your researchPromoSCOPE™ Protein glycosylation is one of the most diverse and abundant forms of post-translational modification in eukaryotic cells, where the attachment of a sugar moieties to protein and lipids increases diversity, to directly regulate cellular functions and intercellular networks, and play important roles in disease and immunity. Glycosylation is involved in many different disease processes and signalling pathways, from the tight control of T-cell development, to its implication in cancer, Alzheimer’s disease, liver diseases and diabetes.Recent developments in single-cell sequencing technologies enable comprehensive characterization of heterogeneous cell populations which provides insights into cellular processes and functions. However, current methods mainly focus on single-cell gene expression profiles, while protein modification is yet to be studied to the same extent at a single-cell resolution.We recently developed PromoSCOPE™, a chemoenzymatic method that quantifies cell surface glycosylation and gene expression profiles in the same single cells. This method utilizes a recombinant enzyme to transfer a DNA-tagged fucose residue to the N-Acetyllactosamine (LacNAc). The tagging DNA contains a polyA tail that can be captured together with mRNA by barcoding beads on Singleron’s SCOPE-chip®. Through sequencing of the DNA tag and mRNA, it is possible to have quantitative multimodal information on both glycosylation and gene expression from the same single cells. Figure 2. Principle of the ProMoSCOPE™ technologyFigure 3. PromoSCOPE™ reveals heterogenous glycosylation abundance in the bone marrow. Single cell suspensions from mouse bone marrow were loaded onto Singleron’s SCOPE-chip® and processed by PromoSCOPE™ Single Cell Glycosylation Detection Kit. (A) UMAP plot showing cell type annotation based on single cell RNAseq results. (B) Tag UMI counts indicating glycosylation abundance and (C) glycosylation abundance in each cell population is shown in the violin plot indicating that immune cells undergo different level of glycosylation.FocuSCOPE®Heterogeneous sample types such as tumor tissues commonly harbor cell-specific genetic variants and gene expression profiles, both of which have been shown to be related to the mechanisms of disease development, progression, and responses to treatment. However, most current high-throughput single cell sequencing methods detect only gene expression levels or epigenetics events such as chromatin conformation. To better decipher the connection between genotype and phenotype at single cell level, we developed FocuSCOPE®, a high-throughput multi-omics sequencing solution that can detect both genetic variants and transcriptome from same single cells.The FocuSCOPE® technology has two key elements: cell-barcoding beads conjugated with oligos that can effectively capture both transcriptome and regions of interest; and reaction chemistry optimized for amplification and library construction of both mRNA sequences and sequences from target regions. FocuSCOPE® has been used to successfully perform single cell analysis of both gene expression profiles and single nucleotide variants, fusion genes, or intracellular viral sequences from thousands of cells simultaneously, delivering more comprehensive insights into each individual cell.Figure 4. Specially designed Barcoding Beads that allow capturing both targeted transcripts and transcriptome.DynaSCOPE®Monitoring RNA dynamics at the single cell level allows us to identify changes that arise from intrinsic properties of biological systems. However, current sequencing technologies provide only a snapshot of the experimental setup. We have overcome this by developing DynaSCOPE® technology which utilizes chemically labeling of nascent mRNA with S4U (4-thiouridine) to ‘timestamp’ all transcripts. During the downstream workflow, S4U is converted to a cytosine analogue. This introduces a mutation that is kept throughout the cDNA amplification and library preparation steps. Only nascent RNAs will contain a mutation which can be bioinformatically detected after sequencing.Figure 5. Schematic representation of the DynaSCOPE® workflow.This method could also be used to investigate mechanisms of infection by bacteria or virus and how individual cells and tissues respond to an antigen or to therapeutic treatment. It can also be applied in vivo by injecting experimental animals with the nucleotide analogue. Thus, it opens the door for more precise understanding of mechanisms of transcription regulation and RNA stability, which might lead to novel therapeutic strategies in the future. Figure 6. (A) UMAP plot of murine lung cells treated (red) and untreated (blue) with the metabolic labeling reagent, shows no differences in the transcriptome. (B) UMAP plot from murine lung tissue from DynaSCOPE® sequencing results. The intensity of the color represents the global level of nascent RNA per cell.Further information about Singleron, these products and other solutions we have to offer is available on our website at www.singleron.bio or contact us via email on info@singleronbio.com and one of our single cell specialists will be happy to talk to you on how we can assist your research. A post by Singleron teamCheck out our latest blog posts Learn more 23.04.03 Linking two worlds: RNA and protein at single cell resolution High-throughput single cell analysis lets researchers see the functions of complex biological systems. Detecting key proteins at the cell surface together with gene expression adds an additional layer oinformation. Read more 23.02.06 Don’t be sO-negative and AB-positive: Donate blood to further hematopoietic research with single cell sequencing Donating blood has the potential to save lives. It can be a lifeline in emergency situations.scRNAseq has recently played a vital role in researching hematological diseases, such as leukemia, multiple myeloma, and lymphoma Read more 23.01.30 Friend or Foe? New Perspectives on Hepatic Immune Cells at Single Cell Resolution Understanding the underlying mechanisms triggering disease development and progression may allow us to identify new diagnostic biomarkers and to develop new treatment strategies. Here, we review recent studies addressing these important questions using singlesequencing technologies. Read more
23.04.03 Linking two worlds: RNA and protein at single cell resolution High-throughput single cell analysis lets researchers see the functions of complex biological systems. Detecting key proteins at the cell surface together with gene expression adds an additional layer oinformation. Read more
23.02.06 Don’t be sO-negative and AB-positive: Donate blood to further hematopoietic research with single cell sequencing Donating blood has the potential to save lives. It can be a lifeline in emergency situations.scRNAseq has recently played a vital role in researching hematological diseases, such as leukemia, multiple myeloma, and lymphoma Read more
23.01.30 Friend or Foe? New Perspectives on Hepatic Immune Cells at Single Cell Resolution Understanding the underlying mechanisms triggering disease development and progression may allow us to identify new diagnostic biomarkers and to develop new treatment strategies. Here, we review recent studies addressing these important questions using singlesequencing technologies. Read more