Features of the organization of bread wheat chromosome 5BS based on physical mapping

Abstract

Background: The IWGSC strategy for construction of the reference sequence of the bread wheat genome is based on first obtaining physical maps of the individual chromosomes. Our aim is to develop and use the physical map for analysis of the organization of the short arm of wheat chromosome 5B (5BS) which bears a number of agronomically important genes, including genes conferring resistance to fungal diseases.

Results: A physical map of the 5BS arm (290 Mbp) was constructed using restriction fingerprinting and LTC software for contig assembly of 43,776 BAC clones. The resulting physical map covered ~ 99% of the 5BS chromosome arm (111 scaffolds, N50 = 3.078 Mb). SSR, ISBP and zipper markers were employed for anchoring the BAC clones, and from these 722 novel markers were developed based on previously obtained data from partial sequencing of 5BS. The markers were mapped using a set of Chinese Spring (CS) deletion lines, and F2 and RICL populations from a cross of CS and CS-5B dicoccoides. Three approaches have been used for anchoring BAC contigs on the 5BS chromosome, including clone-by-clone screening of BACs, GenomeZipper analysis, and comparison of BAC-fingerprints with in silico fingerprinting of 5B pseudomolecules of T. dicoccoides. These approaches allowed us to reach a high level of BAC contig anchoring: 96% of 5BS BAC contigs were located on 5BS. An interesting pattern was revealed in the distribution of contigs along the chromosome. Short contigs (200-999 kb) containing markers for the regions interrupted by tandem repeats, were mainly localized to the 5BS subtelomeric block; whereas the distribution of larger 1000-3500 kb contigs along the chromosome better correlated with the distribution of the regions syntenic to rice, Brachypodium, and sorghum, as detected by the Zipper approach.

Conclusion: The high fingerprinting quality, LTC software and large number of BAC clones selected by the informative markers in screening of the 43,776 clones allowed us to significantly increase the BAC scaffold length when compared with the published physical maps for other wheat chromosomes. The genetic and bioinformatics resources developed in this study provide new possibilities for exploring chromosome organization and for breeding applications.

Keywords: Chromosome 5BS; Genetic markers; Hexaploid wheat; Physical mapping; Sequencing; Synteny; Triticum aestivum.

Fig. 1

Distribution of SSR, ISBP and Zipper markers along the 5B chromosome. Left, genetic map with the distances between markers (cM) and their designations. Right, scheme of the 5BS cytogenetic map; arrows denote boundaries of deletion bins. Bin designations give the information about the arm length they cover; C, centromere. C Banding pattern is given according to http://www.k-state.edu/wgrc/genetic_resources/chinese_spring_deletion_lines.html

Fig. 2

Distribution of assembled contig length. a Distribution of assembly coverage across different size ranges. b Depth of assembly coverage by contig length

Fig. 3

Synteny of wheat (CS) 5BS, wild wheat (T. dicoccoides) 5B, Brachypodium Bd4, rice Os12 and sorghum Sb8, based on BLAST. Selected parts are: 0–322 Mb of dic5B, 0–8.1 Mb of Bd4, 8.9–22.9 of Os12, 34.1–55.4 of Sb8. Wheat 5BS is shown schematically (to demonstrate the distribution of markers along dic5B only: no splitting on resulted physical scaffolds/contigs, position of markers does not exactly correspond to physical position on CS 5BS). Markers from CS BLASTed to T. dicoccoides are based on BAC-end sequences (e-value cutoff 1e-150). Markers to connect dic5B, Bd4 and Os12 are based on CDS sequences of Brachypodium (e-value cutoff 1e-10). Markers to connect Os12 and Sb8 are based on CDS sequences of rice (e-value cutoff 1e-10)

Fig. 4

Distributions of contigs of different lengths on the short arm of 5B chromosome. Contigs were divided into 6 approximately equal groups according to the results of the assembly covered by contigs across different size ranges. Anchoring of BAC contigs to deletion bins was carried out by markers mapped in deletion bins or to 5BS genetic maps