Single-cell whole genome sequencing (scWGS)

We have tested two commercial kits for scWGS library preparation – both based on a 96-well plate format – the Repli-G kit from Qiagen and ResolvomeDNA kit from Bioscryb. We currently offer scWGS with both kits. Further details on the two technologies are provided below.

How to start a scWGS project at NGI

• Contact us at support@ngisweden.se. We’d be happy to set up a meeting to discuss your project!
• Submit an order via the NGI order portal & we’ll get the paperwork set up.
• NGI sends a skirted 96-well plate with the appropriate buffer.
• You sort single cells into the plates and return them to NGI on dry ice.
  • Wells A1 and A2 are reserved for positive and negative controls. 

What we do with your samples

Once at NGI, WGA is done using the protocol of choice. The amplified DNA is then submitted to fragmentation, end-repair and A-tailing followed by adaptor ligation and indexing. In the Bioscryb workflow, the library is also PCR amplified. 

The library is then cleaned and QCed before sequencing. We perform upstream analysis of sequence data including demultiplexing and we provide a Multi-QC report. Data is delivered in the FASTQ format.  

QIAGEN: Multiple Displacement Amplification (MDA)

This method uses a DNA polymerase with strand-displacement activity to amplify DNA with high fidelity and coverage.

Principle of multiple displacement isothermal amplification (MDA): (1) During the annealing step, random MDA hexamer primers bind to the template DNA. (2,3) Strand elongation is achieved using phi29 DNA polymerase with strand displacement activity. (4) The reaction produces a hyperbranched DNA structure. taken from (1). (figure from https://www.qiagen.com).

BioSkryb: Primary Template-directed Amplification (PTA)

PTA is a technique for scWGA that aims to reduce the possible amplification bias produced by MDA and thus improve uniformity (2)

In PTA, a low proportion of termination bases are included in amplification, yielding shorter amplicons compared to a typical MDA reaction. The relatively small size of these amplicons favors subsequent priming of the primary template, thereby limiting the exponential propagation of biases and errors in daughter molecules. (figure from https://www.bioskryb.com/)

References:

1. Volozonoka L, Miskova A, Gailite L. 2022. Whole Genome Amplification in Preimplantation Genetic Testing in the Era of Massively Parallel Sequencing. Int J Mol Sci. Apr 27;23(9):4819. doi: 10.3390/ijms23094819. PMID: 35563216

2. Gonzalez-Pena V, Natarajan S, Xia Y, Klein D, Carter R, Pang Y, Shaner B, Annu K, Putnam D, Chen W, Connelly J, Pruett-Miller S, Chen X, Easton J, Gawad C. 2021. Accurate genomic variant detection in single cells with primary template-directed amplification.Proc Natl Acad Sci U S A. Jun 15;118(24):e2024176118. Doi: 10.1073/pnas.2024176118. PMID: 34099548

Last Updated: 7th May 2025