Taking advantage of evolving and improving sequencing methods, human chromosome 8 is now available as a gapless, end-to-end assembly. Thanks to advances in long-read sequencing technologies, its centromere, telomeres, duplicated gene families and repeat-rich regions are now fully sequenced. We were interested to assess if the new assembly altered our understanding of the potential impact of non-B DNA structures within this completed chromosome sequence. It has been shown that non-B secondary structures, such as G-quadruplexes, hairpins and cruciforms, have important regulatory functions and potential as targeted therapeutics. Therefore, we analysed the presence of putative G-quadruplex forming sequences and inverted repeats in the current human reference genome (GRCh38) and in the new end-to-end assembly of chromosome 8. The comparison revealed that the new assembly contains significantly more inverted repeats and G-quadruplex forming sequences compared to the current reference sequence. This observation can be explained by improved accuracy of the new sequencing methods, particularly in regions that contain extensive repeats of bases, as is preferred by many non-B DNA structures. These results show a significant underestimation of the prevalence of non-B DNA secondary structure in previous assembly versions of the human genome and point to their importance being not fully appreciated. We anticipate that similar observations will occur as the improved sequencing technologies fill in gaps across the genomes of humans and other organisms.

Institute of Biophysics of the Czech Academy of Sciences Královopolská 135 Brno 612 65 Czech Republic

Institute of Biophysics of the Czech Academy of Sciences Královopolská 135 Brno 612 65 Czech Republic; Department of Experimental Biology Faculty of Science Masaryk University Kamenice 5 Brno 62500 Czech Republic

School of Biological Sciences University of East Anglia Norwich Research Park Norwich NR4 7TJ United Kingdom

References provided by Crossref.org

Asymmetric distribution of G-quadruplex forming sequences in genomes of retroviruses

G-quadruplex propensity in H. neanderthalensis, H. sapiens and Denisovans mitochondrial genomes

Interaction of Proteins with Inverted Repeats and Cruciform Structures in Nucleic Acids