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Flow cytometric chromosome sorting from diploid progenitors of bread wheat, T. urartu, Ae. speltoides and Ae. tauschii

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Abstract

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Chromosomes 5A u , 5S and 5D can be isolated from wild progenitors, providing a chromosome-based approach to develop tools for breeding and to study the genome evolution of wheat.

Abstract

The three subgenomes of hexaploid bread wheat originated from Triticum urartu (AuAu), from a species similar to Aegilops speltoides (SS) (progenitor of the B genome), and from Ae. tauschii (DD). Earlier studies indicated the potential of chromosome genomics to assist gene transfer from wild relatives of wheat and discover novel genes for wheat improvement. This study evaluates the potential of flow cytometric chromosome sorting in the diploid progenitors of bread wheat. Flow karyotypes obtained by analysing DAPI-stained chromosomes were characterized and the contents of the chromosome peaks were determined. FISH analysis with repetitive DNA probes proved that chromosomes 5Au, 5S and 5D could be sorted with purities of 78–90 %, while the remaining chromosomes could be sorted in groups of three. Twenty-five conserved orthologous set (COS) markers covering wheat homoeologous chromosome groups 1–7 were used for PCR with DNA amplified from flow-sorted chromosomes and genomic DNA. These assays validated the cytomolecular results as follows: peak I on flow karyotypes contained chromosome groups 1, 4 and 6, peak II represented homoeologous group 5, while peak III consisted of groups 2, 3 and 7. The isolation of individual chromosomes of wild progenitors provides an attractive opportunity to investigate the structure and evolution of the polyploid genome and to deliver tools for wheat improvement.

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Acknowledgments

The authors are grateful to Dr. Jarmila Číhalíková, Bc. Romana Šperková, and Zdenka Dubská for their assistance with chromosome sorting and DNA amplification. This work was funded by the Hungarian National Research Fund (PD83444), by TÁMOP-4.2.2.A-11-1-KONV-2012-0008, by National Science and Technology Office (ALKOBEER OM00363), by János Bolyai Research Scholarships from the Hungarian Academy of Sciences (for MI and CSA), by the Czech Science Foundation (grant award P501/12/G090), by the Ministry of Education, Youth and Sports of the Czech Republic and by the European Regional Development Fund (Operational Programme Research and Development for Innovations No. ED0007/01/01).

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical standards

The authors declare that the experiments comply with the current laws of the countries (Czech Republic and Hungary) in which they were performed.

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Authors and Affiliations

  1. Agricultural Institute, Centre for Agricultural Research, Hungarian Academy of Sciences, Brunszvik u. 2, H-2462, Martonvásár, Hungary

    István Molnár, András Farkas, András Cseh, Mária Megyeri & Márta Molnár-Láng

  2. Centre of the Region Hana for Biotechnological and Agricultural Research, Institute of Experimental Botany, Šlechtitelů 31, 78371, Olomouc, Czech Republic

    Marie Kubaláková, Hana Šimková, Jan Vrána & Jaroslav Doležel

Authors
  1. István Molnár
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  2. Marie Kubaláková
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  3. Hana Šimková
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  4. András Farkas
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  5. András Cseh
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  6. Mária Megyeri
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  7. Jan Vrána
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  8. Márta Molnár-Láng
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  9. Jaroslav Doležel
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Corresponding author

Correspondence to István Molnár.

Additional information

Communicated by B. Friebe.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Online Resource 1. Primer sequences and annealing temperatures of the COS markers used in the present study (DOC 68 kb)

122_2014_2282_MOESM2_ESM.doc

Online Resource 2. PCR products of the COS markers amplified from total genomic DNA (gDNA) of wheat (Mv9kr1), from the diploid progenitors and from subgenomic DNA samples derived from chromosomes sorted from peaks I–III on the flow karyotypes of T. urartu MvGB115, Ae. speltoides MvGB905 and Ae. tauschii MvGB605 (DOC 87 kb)

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Molnár, I., Kubaláková, M., Šimková, H. et al. Flow cytometric chromosome sorting from diploid progenitors of bread wheat, T. urartu, Ae. speltoides and Ae. tauschii . Theor Appl Genet 127, 1091–1104 (2014). https://doi.org/10.1007/s00122-014-2282-2

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  • DOI: https://doi.org/10.1007/s00122-014-2282-2

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