| Autor: |
Nongluk Plongthongkum, Dinh H Diep, Song Chen, Blue Lake, Kun Zhang |
| Rok vydání: |
2020 |
| DOI: |
10.17504/protocols.io.be5gjg3w |
| Popis: |
To study the heterogeneity of complex tissues by joint profiling of gene expression and its regulation, we require an accurate and high-throughput method. Here we described improved high-throughput combinatorial indexing-based single-nucleus chromatin accessibility and mRNA expression sequencing 2 (SNARE-Seq2) co-assay. This protocol involves fixing and permeabilizing the nucleus followed by tagmentation, chromatin barcode ligation, reverse transcription, pooling and splitting for the next rounds of cell barcode ligation into cDNA and accessible chromatin (AC) on the same nucleus. The captured cDNA and AC are co-amplified before splitting and enrichment into single-nucleus RNA and single-nucleus AC sequencing libraries. The protocol can also be applied to both nuclei and whole cells to capture mRNA in the cytoplasm. This improvement allows us to generate hundreds of thousands of data set of each assay and can be scaled up to half a million cells from a single experiment. The entire procedure can be complete in 3.5 d for generating joint single-nucleus RNA and single-nucleus ATAC sequencing libraries. |
| Databáze: |
OpenAIRE |
| Externí odkaz: |
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