DNTR-seq Method Combines WGS, Transcriptomics in Single Cells

Tue, 10/20/2020 - 10:12

2020-10-20 By Quick Biology

Single-cell sequencing is a great method to study cell heterogeneity, track cell lineages. In previous NGS news, we have introduced methods for instances, researchers perform tissue staining combing laser capture microdissection in Geo-seq (Geographical position sequencing) or in situ sequencing in tissue slide sections (slide-seq) to get spatial information of single cells or use metabolically labeled new RNA tagging sequencing to get time-resolved transcripts of single cells. To determine the impact of genomic alterations on gene expression at single-cell resolution, genomic DNA, and mRNA-seq in the same single cell are highly required.

In a paper published in Molecular Cell, scientists in Sweden developed Direct Nuclear Tagmentation and RNA-sequencing (DNTR-seq), which enables whole DNA and transcriptome sequencing jointly in single cells (ref1). Unlike microwell or droplet-based single-cell profiling technology, DNTR-seq does not need specialized equipment. Basically, the protocol lends itself well to a 384-well based format (Fig.1).

1. Cells are firstly deposited using FACs and sorted into plates, allowing enrichment of rare populations by FACS gates.
2. Nuclear and cytosolic compartments are separated by gentle lysis of the cell membrane followed by centrifugation and controlled aspiration. Intact nucleus are in the original sorting plate, while cytosolic is transferred to a new plate.
3. Direct tagmentation of genomic DNA in the nuclear fraction is performed, while the Smart-Seq2 protocol is carried out on the cytosolic fraction.

To test sequencing quality, researchers did their own DNTR-seq, and standard scWGS, Smart-Seq2 protocol using whole-cell extracts. They find DNTR-seq has low position bias in whole genome sequencing and equal mRNA-seq quality compared to RNA-only methods. More independent work should be done to verify DNTR-seq method.

Figure 1: Schematic summary of DNTR-seq (ref1)

Figure 1 Schematic summary of DNTR-seq.png

Quick Biology provides scWGS, scRNA-Seq. Find More at Quick Biology.

Ref:

1. A highly scalable method for joint whole genome sequencing and gene expression profiling of single cells. bioRxiv 2020.03.04 (https://doi.org/10.1101/2020.03.04.976530)