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Telomere-mediated truncation of barley chromosomes

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Abstract

Engineered minichromosomes offer an enormous opportunity to plant biotechnology as they have the potential to simultaneously transfer and stably express multiple genes. Following a top-down approach, we truncated endogenous chromosomes in barley (Hordeum vulgare) by Agrobacterium-mediated transfer of T-DNA constructs containing telomere sequences. Blocks of Arabidopsis-like telomeric repeats were inserted into a binary vector suitable for stable transformation. After transfer of these constructs into immature embryos of diploid and tetraploid barley, chromosome truncation by T-DNA-induced de novo formation of telomeres could be confirmed by fluorescent in situ hybridisation, primer extension telomere repeat amplification and DNA gel blot analysis in regenerated plants. Telomere seeding connected to chromosome truncation was found in tetraploid plants only, indicating that genetic redundancy facilitates recovery of shortened chromosomes. Truncated chromosomes were transmissible in sexual reproduction, but were inherited at rates lower than expected according to Mendelian rules.

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Acknowledgements

We thank Susanne Knüpffer and Cornelia Marthe for excellent technical assistance and Jim Birchler for helpful discussion. This project was supported by the Leibniz Association (WGL) in the context of the ‘Pakt für Forschung und Innovation/WGL Wettbewerb 2009–2011 Leuchtturmprojekte’.

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Authors and Affiliations

  1. Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Corrensstr. 3, 06466, Gatersleben, Germany

    Eszter Kapusi, Lu Ma, Chee How Teo, Götz Hensel, Axel Himmelbach, Ingo Schubert, Michael Florian Mette, Jochen Kumlehn & Andreas Houben

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  1. Eszter Kapusi
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  2. Lu Ma
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  3. Chee How Teo
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  7. Michael Florian Mette
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Correspondence to Andreas Houben.

Additional information

Communicated by Erich Nigg

Eszter Kapusi, Lu Ma and Chee How Teo contributed equally to this work.

Electronic supplementary material

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Supplementary material and method 1

Sequence and annotation of T-DNA constructs (DOC 142 kb)

Supplementary Fig. 1

Transmission of truncated chromosomes in a tetraploid plant via selfing (JPEG 67 kb)

High resolution image (TIFF 79 kb)

Supplementary Fig. 2

Phenotypes of non-transformed tetraploid plants (wild-type, 4n = 28) compared to T0 (BE40/1-1; BE42/2-1) and T1 (BE47/2-11P) plants with variable chromosome numbers, including the presence or absence of truncated chromosomes. Note that plant BE47/2-11P5 shows phenotypical changes without a truncated chromosome, while plant BE47/2-11P6 shows a normal phenotype in the presence of a truncated chromosome (JPEG 113 kb)

High resolution image (TIFF 12017 kb)

Supplementary Table 1

Primers used for vector construction. The introduced restriction sites are indicated in italic (DOC 26 kb)

Supplementary Table 2

Forward and reverse primers and annealing temperature (°C) used for the identification of the various elements of the T-DNA integrated in the genome of barley plants (DOC 27 kb)

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Kapusi, E., Ma, L., Teo, C.H. et al. Telomere-mediated truncation of barley chromosomes. Chromosoma 121, 181–190 (2012). https://doi.org/10.1007/s00412-011-0351-8

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