Chromosomal genomics facilitates fine mapping of a Russian wheat aphid resistance gene

Abstract

Making use of wheat chromosomal resources, we developed 11 gene-associated markers for the region of interest, which allowed reducing gene interval and spanning it by four BAC clones. Positional gene cloning and targeted marker development in bread wheat are hampered by high complexity and polyploidy of its nuclear genome. Aiming to clone a Russian wheat aphid resistance gene Dn2401 located on wheat chromosome arm 7DS, we have developed a strategy overcoming problems due to polyploidy and enabling efficient development of gene-associated markers from the region of interest. We employed information gathered by GenomeZipper, a synteny-based tool combining sequence data of rice, Brachypodium, sorghum and barley, and took advantage of a high-density linkage map of Aegilops tauschii. To ensure genome- and locus-specificity of markers, we made use of survey sequence assemblies of isolated wheat chromosomes 7A, 7B and 7D. Despite the low level of polymorphism of the wheat D subgenome, our approach allowed us to add in an efficient and cost-effective manner 11 new gene-associated markers in the Dn2401 region and narrow down the target interval to 0.83 cM. Screening 7DS-specific BAC library with the flanking markers revealed a contig of four BAC clones that span the Dn2401 region in wheat cultivar 'Chinese Spring'. With the availability of sequence assemblies and GenomeZippers for each of the wheat chromosome arms, the proposed strategy can be applied for focused marker development in any region of the wheat genome.

Accessing a Russian Wheat Aphid Resistance Gene in Bread Wheat by Long‐Read Technologies

“…Five candidate BAC clones, spanning the Dn2401 interval as delimited by Staňková et al, (2015), were Illumina sequenced and assembled, resulting in two scaffolds of 70.8 and 367 kb (Supplemental Fig. S1).…”

“…The 149 newly added individuals of the mapping population (CI2401 × Glupro F 2 –2) were also assayed for the original flanking markers Xowm705 and Xowm711 . Scoring of the RWA response was done at the Colorado State University Insectary in the same fashion as described for the first part of the mapping population (Staňková et al, 2015). Genotype and phenotype data of all 333 individuals from the extended CI2401 × Glupro F 2 mapping population were processed with JoinMap version 4.0 software (Van Ooijen and Voorrips, 2001).…”

“…To identify new markers within the interval delimited by the flanking markers Xowm705 and Xowm711 , we adopted a strategy proposed in our previous study (Staňková et al, 2015) with some modifications. Briefly, to design D‐genome‐specific primers, assembled sequences of five BAC clones spanning the Dn2401 region in 7DS arm of CS were compared with homeologous sequence contigs from the short arms of CS chromosome 7A and 7B (Berkman et al, 2013).…”

Integrated physical map of bread wheat chromosome arm 7DS to facilitate gene cloning and comparative studies

“…Verification of this anchoring approach involved PCR screening of the 7DS-specific BAC library with bread wheat STS markers derived from their Ae. tauschii orthologs [29] and in silico anchoring based on MTP sequence data. The expected positions were confirmed for all STS markers and 460 out of 582 Ae.…”

“…In the current work, we targeted the short arm of bread wheat chromosome 7D (7DS) with the size of 381 Mb [11], which is known to harbor numerous agronomically important genes and QTLs [26][27][28][29]. An early version of a physical map constructed from a 7DS-specific BAC library [30], was used to delimit a region of a gene underlying an aphid resistance [29]. BAC clone Illumina sequences generated in the current study supplemented with OxfordNanopore sequences of a selected BAC clone are being used to identify candidate genes for the pest resistance (Tulpová, unpublished).…”