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bioRxiv · 10.1101/2021.09.24.461328

A method to enrich and purify centromeric DNA from human cells

Gamba, R.·Mazzucco, G.·Wilhelm, T.·Chardon, F.·Velikovsky, L.·Picotto, J.·Doksani, Y.·Fachinetti, D.
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Abstract

Centromeres are key elements for chromosome segregation. Canonical centromeres are built over long-stretches of tandem repetitive arrays. Despite being quite abundant compared to other loci, centromere sequences overall still represent only 2 to 5% of the human genome, therefore studying their genetic and epigenetic features is a major challenge. Furthermore, sequencing of centromeric regions requires high coverage to fully analyze length and sequence variations, which can be extremely costly. To bypass these issues, we have developed a technique based on selective restriction digestion and size fractionation to enrich for centromeric DNA from human cells. Combining enzymes capable of cutting at high frequency throughout the genome, except within most human centromeres, with size-selection of >20 kb fragments resulted in over 25-fold enrichment in centromeric DNA. Sequencing of the enriched fractions revealed that up to 60% of the enriched material is made of centromeric DNA. This approach has great potential for making sequencing of centromeric DNA more affordable and efficient and for single DNA molecule studies.

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Gamba, R., Mazzucco, G., Wilhelm, T., Chardon, F., Velikovsky, L., Picotto, J., Doksani, Y., Fachinetti, D.. 2021-09-24. A method to enrich and purify centromeric DNA from human cells. https://doi.org/10.1101/2021.09.24.461328

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