**Jaroslav Doležel \* , Jana Cˇ ížková , Hana Šimková and Jan Bartoš**

Institute of Experimental Botany, Centre of the Region Haná for Biotechnological and Agricultural Research, Šlechtitel*u*˚ 31, CZ-78371 Olomouc, Czech Republic; cizkova@ueb.cas.cz (J.C.); simkovah@ueb.cas.cz (H.Š); ˇ bartos@ueb.cas.cz (J.B.)

**\*** Correspondence: dolezel@ueb.cas.cz; Tel.: +420-585-238-703; Fax: +420-585-238-704

Received: 26 October 2018; Accepted: 6 November 2018; Published: 11 November 2018

**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.

**Keywords:** flow cytometry; genome size; nuclear DNA content; reference genome assembly; standardization
