As such the smaller genome, with DNA fragments of the same size should still have around 60 linked reads per DNA molecule, but a 10MB genome would mean 5% was in each droplet making the phasing much harder to determine. For example, for a genome whose size is 1/10th the size of the human genome (320 Mb), the mean number of LinkedReads per molecule would be about 6, and the distance between LinkedReads would be about 8 kb, making it hard to anchor barcodes to short initial contigs." My first assumption was that genome size would have no impact on linked read depth, but it would significantly affect the amount of the genome present in a single droplet.
10X CHROMIUM CORE MGH SOFTWARE
It is also fully supported by 10x Genomics CellRanger software run.Question to the authors: I do not understand the statement about smaller genomes getting lower linked read coverage: "For smaller genomes, assuming that the same DNA mass was loaded and that the library was sequenced to the same readdepth, the number of LinkedReads (read pairs) per molecule would drop proportionally, which would reduce the power of the data type. TotalSeq C is specifically designed to be compatible with the 10X Single Cell 5′ workflow used in the Immune Profiling kit.These antibody oligos are complementary to a unique capture sequence on the Single Cell 3′ v3 Gel Beads so antibody barcodes do not compete for the oligo(dT)s capturing the mRNA. TotalSeq B is specifically designed to be compatible with the newer 10X Genomics Single Cell 3’ v3 reagents used in the core and is fully supported by the 10x Genomics CellRanger software.Data must be analyzed using the R based Searat package. At this time, the 10X Genomics analysis pipeline (CellRanger) does not support the processing of the protein derived libraries.
This is the format originally used in the CITE-seq protocol. TotalSeq A is compatible with any Single Cell RNA-seq product that uses poly(dT) for mRNA capture.These are limited to Human and Mouse samples. Antibodies come in three formats that differ in their capture sequence for library preparation so pay close attention to which antibody catalog you use. Detection of multiplets and their removal prior to analysisĪs part of their TotalSeq antibody catalog, BioLegend offers this technology through antibody-oligonucleotide conjugates that can be used for both CITE-seq and Cell Hashing.Increased number of possible replicates in a single experiment.Increased number of cells assayed in a single experiment.Increased sample throughput in a single experiment.Cells assigned a given single tag are binned together, bioinformatically recapitulating the individual samples originally mixed together. Tag assignment enables identification of droplets that originally contained one (singlet) or more cells (multiplets). After cell encapsulation, library preparation, and sequencing, molecular tag information is assigned to cells. The set of multiplexed samples are processed together in a single GEM well. Mostly this comes in two forms – looking for cells that are expressing certain proteins on the cell surface (CITE-seq) or using ubiquitous markers in order to pool different samples together into one channel of the chip (Cell Multiplexing/Cell Hashing).Ĭell or nuclei samples can be labeled with a molecular tag and subsequently mixed with other labeled samples. Single-cell data are analyzed with the Cell Ranger and Loupe Cell Browser software.įeature Barcoding is the broad term used by 10x Genomics in order to describe any method that adds extra layers of information to cells by running single cell gene expression in parallel with other assays to gain useful biological information. Quenching Buffer + Enhancer + 50% Glycerol from 10X Fixation kitġX Nuclei Buffer with RNase Inhibitor and DTT Other standard media with no more than 10% FBS or 2% BSA No EDTA Standard: 10,000 cells per channel up to 80,000 cells per chip Multiplex: 128,000 cells per channel up to 1,024,000 cells per chip Singleplex: 10,000 cells per channel up to 80,000 cells per chip
High: 20,000 cells per channel up to 320,000 cells per chip up to 730,000 cells per chip with plexing Standard: 10,000 cells per channel up to 80,000 cells per chip 140,000 cells per chip with plexing High: 20,000 cells per channel up to 320,000Ĭells per chip up to 730,000 cells per chip with plexing Standard: 10,000 cells per channel up toĨ0,000 cells per chip 140,000 cells per chip Simultaneous detection of mRNA and chromatin accessibilityįixed cells, fixed nuclei, fixed tissue, FFPE tissue Probe-based whole transcriptome gene expression Flavors of 10x Genomics Single Cell Assays
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