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Next generation sequencing analysis reveals a relationship between rDNA unit diversity and locus number in Nicotiana diploids

Roman Matyášek1, Simon Renny-Byfield23, Jaroslav Fulneček1, Jiří Macas4, Marie-Angele Grandbastien5, Richard Nichols2, Andrew Leitch2 and Aleš Kovařík1*

  • * Corresponding author: Aleš Kovařík

  • † Equal contributors

Author Affiliations

1 Institute of Biophysics, Academy of Sciences of the Czech Republic, v.v.i, Královopolská 135, Brno, CZ-612 65, Czech Republic

2 Queen Mary University of London, School of Biological and Chemical Sciences, Mile End Road, London, E1 4NS, UK

3 Department of Ecology, Evolution and Organismal Biology, Iowa State University, Ames, IA 50011, USA

4 Biology Centre, Academy of Sciences of the Czech Republic, Institute of Plant Molecular Biology, Branišovská 31, České Budějovice, CZ-370 05, Czech Republic

5 Institut Jean-Pierre Bourgin, Laboratoire de Biologie Cellulaire, INRA-Centre de Versailles, Versailles Cedex, F-780 26, France

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BMC Genomics 2012, 13:722  doi:10.1186/1471-2164-13-722

Published: 23 December 2012



Tandemly arranged nuclear ribosomal DNA (rDNA), encoding 18S, 5.8S and 26S ribosomal RNA (rRNA), exhibit concerted evolution, a pattern thought to result from the homogenisation of rDNA arrays. However rDNA homogeneity at the single nucleotide polymorphism (SNP) level has not been detailed in organisms with more than a few hundred copies of the rDNA unit. Here we study rDNA complexity in species with arrays consisting of thousands of units.


We examined homogeneity of genic (18S) and non-coding internally transcribed spacer (ITS1) regions of rDNA using Roche 454 and/or Illumina platforms in four angiosperm species, Nicotiana sylvestris, N. tomentosiformis, N. otophora and N. kawakamii. We compared the data with Southern blot hybridisation revealing the structure of intergenic spacer (IGS) sequences and with the number and distribution of rDNA loci.

Results and Conclusions

In all four species the intragenomic homogeneity of the 18S gene was high; a single ribotype makes up over 90% of the genes. However greater variation was observed in the ITS1 region, particularly in species with two or more rDNA loci, where >55% of rDNA units were a single ribotype, with the second most abundant variant accounted for >18% of units. IGS heterogeneity was high in all species. The increased number of ribotypes in ITS1 compared with 18S sequences may reflect rounds of incomplete homogenisation with strong selection for functional genic regions and relaxed selection on ITS1 variants. The relationship between the number of ITS1 ribotypes and the number of rDNA loci leads us to propose that rDNA evolution and complexity is influenced by locus number and/or amplification of orphaned rDNA units at new chromosomal locations.

rRNA genes; Concerted evolution; Plants; Chromosomes; DNA repeats homogenisation