Prospects of telomere‐to‐telomere assembly in barley: Analysis of sequence gaps in the MorexV3 reference genome

Abstract

The first gapless, telomere-to-telomere (T2T) sequence assemblies of plant chromosomes were reported recently. However, sequence assemblies of most plant genomes remain fragmented. Only recent breakthroughs in accurate long-read sequencing have made it possible to achieve highly contiguous sequence assemblies with a few tens of contigs per chromosome, that is a number small enough to allow for a systematic inquiry into the causes of the remaining sequence gaps and the approaches and resources needed to close them. Here, we analyse sequence gaps in the current reference genome sequence of barley cv. Morex (MorexV3). Optical map and sequence raw data, complemented by ChIP-seq data for centromeric histone variant CENH3, were used to estimate the abundance of centromeric, ribosomal DNA, and subtelomeric repeats in the barley genome. These estimates were compared with copy numbers in the MorexV3 pseudomolecule sequence. We found that almost all centromeric sequences and 45S ribosomal DNA repeat arrays were absent from the MorexV3 pseudomolecules and that the majority of sequence gaps can be attributed to assembly breakdown in long stretches of satellite repeats. However, missing sequences cannot fully account for the difference between assembly size and flow cytometric genome size estimates. We discuss the prospects of gap closure with ultra-long sequence reads.

Telomere-to-telomere and gap-free reference genome assembly of the kiwifruit Actinidia chinensis

“…This helped us to achieve the completely gap-free kiwifruit reference genome Hongyang v4.0. Based on our results, it is recommended to use the HiFi contigs assembled by hifiasm [ 13 ] as a fixed backbone and then perform manual handling with the ONT contigs to fill the small amount of remnant gaps, which is identical to the genome assembly strategies adopted in rice, watermelon, and barley [ 10 , 32 , 33 ], but slightly different from those in Arabidopsis and banana [ 8 , 11 ]. Comprehensively, the choice of genome assembly strategy should depend on practical testing effects, which may vary among different species.…”

A technical guide to TRITEX, a computational pipeline for chromosome-scale sequence assembly of plant genomes

“…We anticipate that manual curation of genome assemblies will be needed for at least the next 3 to 5 years to construct and validate chromosomal pseudomolecules. Navrátilová et al [ 9 ] have singled out one factor that can prevent telomere-to-telomere assembly in complex plant genomes: long homogenous satellite arrays with nucleotide compositions unfavorable to the PacBio platform. New technologies may overcome these obstacles.…”

“…Strategies and tools to obtain haplotype-resolved assemblies have been reviewed in detail elsewhere [ 7 , 8 ]. The assembly even of long and accurate sequence reads rarely results in sequence contigs spanning entire chromosomes from telomere to telomere [ 9 ]. Complementary linkage information is needed to arrange contigs into chromosome-scale scaffolds.…”

A technical guide to TRITEX, a computational pipeline for chromosome-scale sequence assembly of plant genomes

“…We anticipate that manual curation of genome assemblies will be needed for at least the next three to ve years to construct and validate chromosomal pseudomolecules. Navrátilová et al (7) have singled out one factor that can prevent telomere-to-telomere assembly in complex plant genomes: long homogenous satellite arrays with nucleotide compositions unfavorable to the PacBio platform. New technologies may overcome these obstacles.…”