The 5S intergenic spacer
The 5S rRNA genes form gene arrays analogous to the 35S rDNA, localized
on one or a few chromosomes, usually afar from the nucleolus organizer
regions (NORs) of the 35S rDNA
(http://www.plantrdnadatabase.com/;
see Ribeiro et al. 2011 for selected oak and beech species). In higher
plants, the number of repeats per genome ranges from hundreds to
thousands (Cloix et al. 2000), and the 5S rRNA coding regions are
separated by a generally variable, non-transcribed intergenic spacer
(5S-IGS) with high phylogenetic resolution (e.g., Forest et al. 2005;
Blattner et al. 2009; Grimm and Denk 2010; Garcia and Kovařik 2013;
Mlinarec et al. 2016). Its inter-individual and intra-genomic
variability is probably the main reason why it is little used. It cannot
be directly sequenced but requires cloning and methodological frameworks
that can make use of the often higher intra-genomic than
inter-individual diversity (Göker and Grimm 2008; Potts et al. 2014; for
an application using 5S-IGS data see e.g. Simeone et al. 2018).
Therefore, large numbers of 5S-IGS variants should ideally be cloned and
sequenced to identify intra- and inter-array polymorphism, and
phylogenetic signals useful for inferring evolutionary patterns (Eidesen
et al. 2007). In previous studies, Denk and Grimm (2010) and Simeone et
al. (2018) produced a data set with more than 1000 Sanger-sequenced
5S-IGS clones (covering 30 species, nearly 300 individuals) and
demonstrated the potential of this marker for the circumscription of
most of the investigated oak species, to recognize hybridization, and to
infer reticulation/introgression events.