Wild wheat relative Aegilops biuncialis offers valuable traits for crop improvement through interspecific hybridization. However, gene transfer from Aegilops has been hampered by difficulties in detecting introgressed Ub- and Mb-genome chromatin in the wheat background at high resolution. The present study applied DArTseq technology to genotype two backcrossed populations (BC382, BC642) derived from crosses of wheat line Mv9kr1 with Ae. biuncialis accession, MvGB382 (early flowering and drought-tolerant) and MvGB642 (leaf rust-resistant). A total of 11,952 Aegilops-specific Silico-DArT markers and 8,998 wheat-specific markers were identified. Of these, 7,686 markers were assigned to Ub-genome chromosomes and 4,266 to Mb-genome chromosomes and were ordered using chromosome scale reference assemblies of hexaploid wheat and Ae. umbellulata. Ub-genome chromatin was detected in 5.7% of BC382 and 22.7% of BC642 lines, while 88.5% of BC382 and 84% of BC642 lines contained Mb-genome chromatin, predominantly the chromosomes 4Mb and 5Mb. The presence of alien chromatin was confirmed by microscopic analysis of mitotic metaphase cells using GISH and FISH, which allowed precise determination of the size and position of the introgression events. New Mv9kr1-Ae. biuncialis MvGB382 4Mb and 5Mb disomic addition lines together with a 5DS.5DL-5MbL recombination were identified. A possible effect of the 5MbL distal region on seed length has also been observed. Moreover, previously developed Mv9kr1-MvGB642 introgression lines were more precisely characterized. The newly developed cytogenetic stocks represent valuable genetic resources for wheat improvement, highlighting the importance of utilizing diverse genetic materials to enhance wheat breeding strategies.

Agricultural Research Centre Field Crops Research Institute 9 Gamma Street Giza 12619 Egypt

Department of Biological Resources Centre for Agricultural Research Hungarian Research Network Martonvásár 2462 Hungary

Institute of Experimental Botany of the Czech Academy of Sciences Centre of Plant Structural and Functional Genomics Olomouc 77900 Czech Republic

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