Flow cytometry to characterize nuclear genome and chromosome sorting of Agropyron cristatum, a wild relative of wheat

Said M., Vrána J., Karafiátová M., Čížková J., Cabrera A., Doležel J.
International Conference Analytical Cytometry IX October 14 - 17 Prague Czech Republic : , 2017

Abstract: Crested wheatgrass (Agropyron cristatum L. Gaertn.) is a wild relative of wheat which exhibits large genetic variation and offers attractive source of novel genes for wheat improvement. The species comprises a complex of diploid (2n=2x=14, PP), tetraploid (2n=4x=28, PPPP) and hexaploid (2n=6x=42, PPPPPP) forms. In the first part of this work, we characterized 80 accessions collected from different parts of the world. Flow cytometric analysis classified 26 accessions as diploid, 51 accessions as tetraploid and 3 accessions as hexaploid. These results were confirmed by mitotic cell chromosome counts. Mean nuclear 2C DNA content of diploid plants was estimated as 12.83 pg DNA (1C genome size = 6.27 Gbp). Some diploid genotypes exhibited slightly higher DNA content, and were identified by FISH analysis as B-chromosome carriers. With the aim to facilitate genome mapping and sequencing, we explored a possibility of developing flow cytogenetics for the species. Flow karyotypes obtained after the analysis of DAPI-stained suspensions of mitotic metaphase chromosomes of diploid A. cristatum consisted of three peaks representing the seven chromosomes pairs of the species. The chromosomes could only be sorted in groups of 2 or 3. However, chromosomes 1P - 6P could be sorted individually from wheat-A. cristatum addition lines at purities ranging from 81 to 98%. B chromosomes could be easily discriminated and sorted at purities higher than 95%. These results provide an attractive opportunity to investigate the structure and evolution of the P genome of A. cristatum and to develop molecular tools to facilitate the exploitation of this wild species and support alien introgression breeding of bread wheat. This work has been supported by the Grant Agency of the Czech Republic (award 315160-2015).
IEB authors: Mahmoud Said, Jana Čížková, Jaroslav Doležel, Miroslava Karafiá..., Jan Vrána