![]() The 10X Genome sequencing data contain conventional shotgun sequencing data, after trimming off the first 26 bases of the forward read. For human whole genome phasing and structural variant analysis GemCode sequencing data at 60x genome coverage or a combination of conventional Illumina data coverage at 30x genome coverage and GemCode at least 23x is recommended. Human phasing can be carried out using both whole genome shotgun sequencing as well as employing exome capture enrichment. A single HiSeq lane generates sufficient information for phasing and structural variant analysis of a human sized genome. Sub-genome sized parts of the DNA sample (about 1/6000th of a genome) are compartmentalized in nanoliter volume oil droplets together with beads carrying one of about two million different barcodes to initiate the library preparation with droplet specific barcodes. Please note that the individual DNA molecules are not meant to be fully assembled with this approach. ![]() DNA equivalent to about 150 copies of a genome does get distributed over about 1 million oil droplets that include beads. Thus, it allows individual long DNA molecules 10X Genomics Chromium Genome Linked-reads principle. The 10X Genomics technology generates individually barcoded sequencing libraries for hundreds of thousands of nanoliter volume oil droplets using up to 1.7 million different barcodes. Please see these slides for details on GemCode technology and some of the possible applications. Genotyping in repetitive regions of genomes is possible via an approach called “critical content rescue”. The applications currently supported by the 10X Genomics software are human haplotype phasing and structural variant detection (see this page for currently supported applications) as well as de novo genome assemblies (please see below). Just a few nanograms of sample ar e required as input for the library preparation. When analyzing high molecular weight DNA samples the linked read sequencing data delineate linkage information over distances of up to 150 kb. The sequencing libraries are generated with the Chromium controller instrument and reagents from 10X Genomics, followed by sequencing on our Illumina HiSeq sequencers. We are offering 10X Genomics Chromium Genome “linked read” library preps and sequencing. Please contact to reserve reagents for your project. As soon as these run out, we will discontinue the assay effective immediately. Update July 2020: We have ~16 remaining 10X Chromium Genome reactions left in our core. ![]() ![]() We are sad to see this very popular assay go. ![]() Libraries were quantified using Agilent Bioanalyzer High Sensitivity DNA chips and pooled together to get similar numbers of reads from each single cell before sequencing on the NovaSeq S4 (by Fulgent Genomics or Novogene).As of January 2020, 10X Genomics has announced that it is discontinuing production of the 10X Chromium Genome linked read reagents. The single cell 3' gene expression libraries were constructed and cDNA corresponding to an insertion size of around 350 bp selected. After GEM-RT and cleanup, the cDNA from barcoded single cell RNAs were amplified before quantification using Agilent Bioanalyzer High Sensitivity DNA chips. Briefly, Gel Bead-In EMulsions (GEMs) are generated using single cell preparations. scRNA-seq libraries were prepared following the instructions from Chromium Single Cell 3' Reagent Kits User Guide (v2 or v3). Single cells are captured and barcoded using the 10X Chromium platform (10X Genomics). We profiled transcriptomes of >370,000 individual cells from endometriomas (n=8), endometriosis (n=28), eutopic endometrium (n=10), unaffected ovary (n=4) and endometriosis-free peritoneum (n=4) to create a cellular atlas of endometrial-type epithelial cells, stromal cells, and microenvironmental cell populations across tissue sites. GEO help: Mouse over screen elements for information.Ī single-cell transcriptomic analysis of endometriosisĮxpression profiling by high throughput sequencing ![]()
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