de novo re-construction of the core genome from ChIP-Seq for large-genome organisms

Genetic diversity in plants is remarkably high. Recent whole genome sequencing (WGS) of 67 rice accessions recovered 10,872 novel genes. Comparison of the genetic architecture among divergent populations or between crops and wild relatives is essential for obtaining functional components determining crucial traits. However, many major crops have gigabase-scale genomes, which are not well-suited to WGS. Existing cost-effective sequencing approaches including re-sequencing, exome-sequencing and restriction enzyme-based methods all have difficulty in obtaining long novel genomic sequences from highly divergent population with large genome size. The present study presented a reference-independent core genome targeted sequencing approach, CGT-seq, which employed epigenomic information from both active and repressive epigenetic marks to guide the assembly of the core genome mainly composed of promoter and intragenic regions. This method was relatively easily implemented, and displayed high accuracy, sensitivity and specificity for capturing the core genome of bread wheat. 95% intragenic and 89% promoter region from wheat were covered by CGT-seq read. We further demonstrated in rice that CGT-seq captured hundreds of novel genes and regulatory sequences from a previously unsequenced ecotype. Together, with specific enrichment and sequencing of regions within and nearby genes, CGT-seq is a time- and resource-effective approach to profiling functionally relevant regions in sequenced and non-sequenced populations with large genomes. We designed an experimental and computational pipeline for re-construction of wheat core genome. Briefly, ChIP-seq of different marks was performed, sequencing reads of each modification were assembled and further merged. Next, reads from all modifications were mapped to the assembled contigs derived from the previous step, and paired sequencing reads were used to join contigs into scaffolds. All reads were re-mapped to the scaffolds, gaps were filled and scaffolds were extended. The scaffolding and extension steps were repeated until the covered bases showed no apparent change.