Illumina Library Preparation for Sequencing the GC-Rich Fraction of Heterogeneous Genomic DNA

Marie-Ka Tilak (1) , Fidel Botero-Castro (1) , Nicolas Galtier (2) , Benoit Nabholz (1)

1 UMR ISEM - Institut des Sciences de l'Evolution de Montpellier
2 EPHE - École Pratique des Hautes Études

Standard Illumina libraries are biased toward sequences of intermediate GC-content. This results in an underrepresentation of GC-rich regions in sequencing projects of genomes with heterogeneous base composition, such as mammals and birds. We developed a simple, cost-effective protocol to enrich sheared genomic DNA in its GC-rich fraction by subtracting AT-rich DNA. This was achieved by heating DNA up to 90 C before applying Illumina library preparation. We tested the new approach on chicken DNA and found that heated DNA increased average coverage in the GC-richest chromosomes by a factor up to six. Using a Taq polymerase supposedly appropriate for PCR amplification of GC-rich sequences had a much weaker effect. Our protocol should greatly facilitate sequencing and resequencing of the GC-richest regions of heterogeneous genomes, in combination with standard short-read and long-read technologies.

Mots clés: GC content; GC enrichment; high-throughput sequencing; bird

Domaines: Génétique des populations [q-bio.PE]; Systématique, phylogénie et taxonomie; Génomique, Transcriptomique et Protéomique [q-bio.GN]

Marie-Ka Tilak, Fidel Botero-Castro, Nicolas Galtier, Benoit Nabholz. Illumina Library Preparation for Sequencing the GC-Rich Fraction of Heterogeneous Genomic DNA. Genome Biology and Evolution, 2018, 10 (2), pp.616-622. ⟨10.1093/gbe/evy022⟩. ⟨hal-01814697⟩