Abstract
Any project seeking to deliver a plant or animal reference genome sequence must address the question as to the completeness of the assembly. Given the complexity introduced particularly by the presence of sequence redundancy, a problem which is especially acute in polyploid genomes, this question is not an easy one to answer. One approach is to use the sequence data, along with the appropriate computational tools, the other is to compare the estimate of genome size with an experimentally measured mass of nuclear DNA. The latter requires a reference standard in order to provide a robust relationship between the two independent measurements of genome size. Here, the proposal is to choose the human male leucocyte genome for this standard: its 1C DNA amount (the amount of DNA contained within unreplicated haploid chromosome set) of 3.50 pg is equivalent to a genome length of 3.423 Gbp, a size which is just 5% longer than predicted by the most current human genome assembly. Adopting this standard, this paper assesses the completeness of the reference genome assemblies of the leading cereal crops species wheat, barley and rye.
Highlights
The more that is known regarding the organization and function of plant and animal genomes, the more it becomes clear that a full understanding of genome function will require the acquisition of a complete sequence
The level of completeness of an assembly remains difficult to ascertain, especially in the case of complex genomes, in which tracts of repetitive DNA, segmental duplications and, in the case of polyploid genomes, the presence of homoeologs, are all inimical to the elaboration of a “correct” assembly: the result is that gaps, mis-assemblies and collapsed tandem repeats feature in most published genome sequences
To enable a comparison of data obtained by different laboratories, Tiersch et al [9] calibrated a set of animal reference standards, choosing human male leukocytes (7 pg DNA/2C) as the primary reference; the 7 pg figure was based on estimates derived from Feulgen micro-densitometry [10]
Summary
The more that is known regarding the organization and function of plant and animal genomes, the more it becomes clear that a full understanding of genome function will require the acquisition of a complete sequence. The enormous throughput offered by current short read DNA sequencing technologies allows for the sequencing of genomes of any size and at a high sequencing depth. While this enables the ready assembly of single and low-copy sequences, the inclusion within the assembly of repetitive sequence is a non-trivial challenge, and, together with sequence redundancy due to polyploidy, represent a major obstacle to the acquisition of gap-free long-range genome sequences. Genome size of unknown species might be obtained by extrapolation, using data from species whose genome size is known As both approaches rely on sequence data, the only truly independent way to determine genome size is to experimentally determine the quantity of DNA present in the nuclei
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