Next Generation Arabidopsis Genomics

The Arabidopsis 1,001-Epigenomes project provides an unprecedented resource to discover transcriptomes and methylomes for over 1,000 natural A. thaliana accessions. BioNano Genomics optical genome maps for nine divergent accessions added an additional layer that enabled the identification and scaffolding for over 600 highly diverse genome regions per accession. These variations create balanced gains and losses of over 3 Mb between accessions, and provided evidence for TE-rich disease resistance gene loci being the most divergent. To further analyze and understand the content of structural variations, we have created the highest quality reference genomes for six diverse A. thaliana accessions. PacBio RSII sequencing in combination with optical genome maps and Illumina PCR-free libraries enabled us to assemble, correct and annotate complete chromosome arms. RNA-seq, as well as whole genome bisulfite sequencing, enabled the description of the transcriptome and methylome for four tissue types, which provided unprecedented insight into highly divergent regions of structural variations.

Previously it has been demonstrated that transgene insertions also result in smaller structural variations. To study the effects of T-DNA insertion on genome structure, we created optical genome maps for five individual mutant lines from the SALK, the WiscSeq and SAIL collections. These T-DNA insertion lines represent a collection of over 200,000 mutant A. thaliana Col-0 lines and are an important resource to study gene function. While the transformation binary vectors are approximately 13 kb in size, we identified T-DNA insertion sites ranging from 28 kb to 206 kb, consistent with multiple concatenated insertion events. To further investigate sequence content, we utilized long read Oxford Nanopore MinION sequencing and confirmed tandem arrays of T-DNA plus vector backbone sequences within insertions. Identification of such large insert sizes and potential immediate CG methylation of transgenes will have implications for how we create and analyze transgenic events in the future.