Whole genome amplification (WGA) increases DNA amount in very limited samples to meet high input needs of downstream analyses like sequencing. Among other methods, multiple displacement amplification (MDA) has emerged as a relatively uncomplicated WGA method that uses random hexamer primers to amplify with high fidelity all DNA sequences in a sample.
Though touted to be accurate, biases, allelic dropout, and intermolecular chimeric products are known issues of MDA in a tube. The droplet MDA (dMDA) used in the Xdrop technology resolves these by encapsulating single-molecule amplification reactions in droplets.
Xdrop dMDA of DNA from single cells outperforms standard MDA and provides uniform coverage across the genome. Our collaborators, Joanna Hård, Jeff Mold, Jakob Michaëlsson from Karolinska Institutet and Adam Ameur from SciLife Lab Uppsala University, studied the performance of whole genome amplification on compartmentalized single DNA molecules (dMDA) versus standard MDA in recovering genomic information from single human cells.
Visible in the graph (right), is the average read depth per chromosome from Illumina whole genome sequencing of single cells amplified with Xdrop dMDA (red), standard MDA (green) and an unamplified bulk sample (black). Xdrop dMDA achieves uniform genome wide coverage that is comparable to an unamplified bulk sample. In contrast, standard MDA exhibits uneven coverage due to amplification bias.
We compared the performance of Xdrop droplet MDA (dMDA) to alternative WGA solutions based on standard MDA in a tube. We used each solution to amplify serially diluted Escherichia coli DNA, from 0 (no-template control) to 1000 pg. Xdrop performed exceptionally well across the entire input range but exceled where the standard MDA methods reached their limits. Xdrop was also the only method to amplify 1 pg input DNA and the output covered 99% of the E. coli genome. View the complete dataset in our Application Note below.
Especially noteworthy is the extraordinary uniformity in genome coverage we achieved with Xdrop.
Each molecule in a droplet was amplified individually and thus, when we pooled the contents to generate the sequencing library, all DNA molecules of the sample were represented in true proportions in the sequencing results. This was not the case for the standard MDA solutions in a tube, where coverage was extremely variable and biased.
We corroborated the amplification efficiency of Xdrop dMDA with sub-picogram amounts of human DNA. Check out our scientific poster below for the details.
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