Nuclei or Chromatin Isolation from Single-Cell Types
Capture and isolation of single cells are prerequisites for understanding these cellular variations. However, efficient single-cell capture and isolation are complex tasks for any cell population. For the interrogation of chromatin, nuclei are sometimes preferred over whole cells to avoid cytoplasm contamination. In addition, nuclei can be cross-linked more feasibly than whole cells. For these reasons, nuclei-based methods have been developed.
Isolation of Nuclei in Tagged Cell Types (INTACT)
INTACT is a method applicable for the epigenetic profiling and gene expression analysis pattern of almost all model organisms. Cell type-specific nuclei have been isolated using the tag of NTF (nuclear targeting fusion) protein. NTF tag comprises three parts: a nuclear envelope localizing protein, a part of green fluorescent protein (GFP) protein for visualization, and the biotin acceptor peptide BLRP (biotin ligase recognition peptide). Biotinylated nuclei can be sorted using streptavidin magnetic beads.
Fig.1 INTACT method diagram. (Deal, 2010)
Advantages of INTACT
- High specificity.
- Cost-effective.
- Easy to apply, no fixation.
Limitations of INTACT
- Time-consuming for the production of genetic tags.
- Low throughput.
Batch Isolation Tissue-Specific Chromatin for Immunoprecipitation (BiTS-ChIP)
BiTS-ChIP is a method for isolating tissue-specific chromatin, and it was introduced to obtain nuclei from fixed embryos by using fluorescence Activated Cell Sorting (FACS). It is a cell type-specific ChIP technique based on the tagging of nuclear protein expressed in a specific tissue.
Fig.2 BiTS-ChIP method diagram. (Bonn, 2012)
Advantages of BiTS-ChIP
- Time-effective.
- Excellent purity.
Limitations of BiTS-ChIP
- Time-consuming for the production of genetic tags.
- Antibody-based sorting method, which may make it expensive.
Nuclear Tagging and Translating Ribosome Affinity Purification (NuTRAP)
The nuTRAP method combines cell type-specific isolation from both nuclei and ribosomes. Similar to INTACT, the nuclear membrane is labeled by a fusion protein during the NuTRAP. And the NuTRAP ribosome is captured by the TRAP (Translating Ribosome Affinity Purification) method.
The nuTRAP method was used to investigate the in vivo chromatin states to reveal the epigenomics of adipocytes. It is a highly efficient method that enables more accessible access to diverse cell types, which is challenging for in vitro ChIP-seq.
Fig.3 Scheme of NuTRAP mouse. (Roh, 2017)
Advantages of NuTRAP
- High specificity.
- Cost-effective
- No fixation.
Limitations of NuTRAP
- Time-consuming for the production of genetic tags.
- Low throughput.
At Creative Biolabs, our professional team provides one-stop nuclei or chromatin isolation from single-cell types services including INTACT, BiTS-ChIP, and NuTRAP for a better solution for single-cell nuclei or chromatin analysis. If you have any requirements, please feel free to contact us for further communication about your project.
References
- Dean, R.B.; et al. The INTACT method for cell type-specific gene expression and chromatin profiling in Arabidopsis thaliana. Nature Protocols. 2010, 6: 56.
- Bonn, S.; et al. Tissue-specific chromatin immunoprecipitation from multicellular complex samples using BiTS-ChIP. Nature Protocols. 2012, 7: 978-994.
- Roh, H.C.; et al. Simultaneous transcriptional and epigenomic profiling from specific cell types within heterogeneous tissues in vivo. Cell Reports. 2017, 18: 1048-1061.
