The isolation of nuclei is achieved by 10× Genomics ChromiumTM, which consists eight-channel microfluidics system with double crossings. In this system, a gel beads with barcodes and primer, enzymes and a single nucleus are encapsulated in nanoliter-sized oil drop, generating Gel Bead-in-Emulsion (GEM). Once GEM are formed, cell lysis and release of barcodes are performed in each GEM. mRNA are reverse transcribed into cDNA molecules with 10× barcodes and UMI, which are further subject to standard sequencing library construction.
Cell / Tissue |
Reason |
Unfresh frozen tissue |
Unable to get fresh or long-saved organizations |
Muscle cell, Megakaryocyte, Fat… |
Cell diameter is too large to enter the instrument |
Liver… |
Too fragile to break, unable to distinguish single cells |
Neuron cell, Brain… |
More sensitive, easy to stress, will change the sequencing results |
Pancreas, Thyroid… |
Rich in endogenous enzymes, affecting the production of single cell suspension |
Single-nucleus |
Single-cell |
Unlimited cell diameter |
Cell diameter: 10-40 μm |
The material can be frozen tissue |
The material must be fresh tissue |
Low stress of frozen cells |
Enzyme treatment may cause cell stress reaction |
No red blood cells need to be removed |
Red blood cells need to be removed |
Nuclear expresses bioinformation |
The whole cell expresses bioinformation |
Library |
Sequencing strategy |
Data Volume |
Sample Requirements |
Tissue |
10× Genomics single-nuclei library |
10x Genomics -Illumina PE150 |
100,000 reads/cell approx. 100-200 Gb |
Cell number: >2× 105 Cell conc. at 700-1,200 cell/μL |
≥ 200 mg |
For more details on sample preparation guidance and service workflow, please feel free to talk to a BMKGENE expert
1.Spot clustering
2.Marker expression abundance clustering heatmap
3.Maker gene distribution in different clusters
4.Cell trajectory analysis/pseudotime