High-throughput single-cell epigenomic profiling by targeted insertion of promoters (TIP-seq)
| Autor: | David M. Gilbert, Vishnu Dileep, Tetsuya Handa, Daniel A. Bartlett, Hiroshi Kimura, Steven Henikoff, Yasuyuki Ohkawa |
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| Rok vydání: | 2021 |
| Předmět: |
Epigenomics
biology Chromosome Mapping RNA polymerase II Cell Biology Computational biology HCT116 Cells Chromatin DNA binding site Mutagenesis Insertional chemistry.chemical_compound chemistry biology.protein medicine Humans T7 RNA polymerase Single-Cell Analysis Promoter Regions Genetic Transcription factor Polymerase DNA medicine.drug |
| Zdroj: | Journal of Cell Biology. 220 |
| ISSN: | 1540-8140 0021-9525 |
| Popis: | Chromatin profiling in single cells has been extremely challenging and almost exclusively limited to histone proteins. In cases where single-cell methods have shown promise, many require highly specialized equipment or cell type–specific protocols and are relatively low throughput. Here, we combine the advantages of tagmentation, linear amplification, and combinatorial indexing to produce a high-throughput single-cell DNA binding site mapping method that is simple, inexpensive, and capable of multiplexing several independent samples per experiment. Targeted insertion of promoters sequencing (TIP-seq) uses Tn5 fused to proteinA to insert a T7 RNA polymerase promoter adjacent to a chromatin protein of interest. Linear amplification of flanking DNA with T7 polymerase before sequencing library preparation provides ∼10-fold higher unique reads per single cell compared with other methods. We applied TIP-seq to map histone modifications, RNA polymerase II (RNAPII), and transcription factor CTCF binding sites in single human and mouse cells. |
| Databáze: | OpenAIRE |
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