Remarkable variation of ribosomal DNA organization in Gnetophytes, a distinct lineage of gymnosperms

Gnetophytes, comprising genera Ephedra, Gnetum and Welwitschia, are an understudied, enigmatic lineage of gymnosperms, whose phylogenetic relationships with other seed plants remains controversial. Here we examine the organization of ribosomal DNA (rDNA) across representative species. Analysing low-coverage whole genome sequencing (NGS) data in combination with applying the conventional fluorescence In Situ Hybridization (FISH) and Southern blot hybridization techniques we found that: 1) rDNA in three gnetophyte species have distinctive rDNA unit organisation, 2) Length of 35S and 5S rDNA units varies considerably,3) 5S rDNA of G. montanum has high genomic heterogeneity compared to 35S rDNA or 5S rDNAs in other two species, which suggests lower frequencies of 5S rDNA homogenization and/or pseudogenisation in G. montnanum. 4) 5S rDNA in W. mirabilis forms three unique clusters in the genome and has surprisingly low genomic copy numbers and heterogeneity, indicating efficient, relatively high frequency of concerted evolution.