title: Exploring the tertiary gene pool of bread wheat: sequence assembly and analysis of chromosome 5M(g) of Aegilops geniculata.
abstract: Next-generation sequencing (NGS) provides a powerful tool for the discovery of important genes and alleles in crop plants and their wild relatives. Despite great advances in NGS technologies, whole-genome shotgun sequencing is cost-prohibitive for species with complex genomes. An attractive option is to reduce genome complexity to a single chromosome prior to sequencing. This work describes a strategy for studying the genomes of distant wild relatives of wheat by isolating single chromosomes from addition or substitution lines, followed by chromosome sorting using flow cytometry and sequencing of chromosomal DNA by NGS technology. We flow-sorted chromosome 5M(g) from a wheat/Aegilops geniculata disomic substitution line [DS5M(g) (5D)] and sequenced it using an Illumina HiSeq 2000 system at approximately 50 × coverage. Paired-end sequences were assembled and used for structural and functional annotation. A total of 4236 genes were annotated on 5M(g) , in close agreement with the predicted number of genes on wheat chromosome 5D (4286). Single-gene FISH indicated no major chromosomal rearrangements between chromosomes 5M(g) and 5D. Comparing chromosome 5M(g) with model grass genomes identified synteny blocks in Brachypodium distachyon, rice (Oryza sativa), sorghum (Sorghum bicolor) and barley (Hordeum vulgare). Chromosome 5M(g) -specific SNPs and cytogenetic probe-based resources were developed and validated. Deletion bin-mapped and ordered 5M(g) SNP markers will be useful to track 5M-specific introgressions and translocations. This study provides a detailed sequence-based analysis of the composition of a chromosome from a distant wild relative of bread wheat, and opens up opportunities to develop genomic resources for wild germplasm to facilitate crop improvement.
authors.affiliation: Department of Plant Pathology, Wheat Genetics Resource Center, Kansas State University Manhattan, Manhattan, KS, 66506, USA.
authors.forename: Vijay K
authors.initials: VK
authors.lastname: Tiwari
authors.affiliation: Department of Plant Pathology, Kansas State University, Manhattan, KS, 66502, USA.
authors.forename: Shichen
authors.initials: S
authors.lastname: Wang
authors.affiliation: Department of Plant Pathology, Wheat Genetics Resource Center, Kansas State University Manhattan, Manhattan, KS, 66506, USA.
authors.forename: Tatiana
authors.initials: T
authors.lastname: Danilova
authors.affiliation: Department of Plant Pathology, Wheat Genetics Resource Center, Kansas State University Manhattan, Manhattan, KS, 66506, USA.
authors.forename: Dal Hoe
authors.initials: DH
authors.lastname: Koo
authors.affiliation: Institute of Experimental Botany, Centre of the Region Haná for Biotechnological and Agricultural Research, CZ 78371, Olomouc, Czech Republic.
authors.forename: Jan
authors.initials: J
authors.lastname: Vrána
authors.affiliation: Institute of Experimental Botany, Centre of the Region Haná for Biotechnological and Agricultural Research, CZ 78371, Olomouc, Czech Republic.
authors.forename: Marie
authors.initials: M
authors.lastname: Kubaláková
authors.affiliation: Institute of Experimental Botany, Centre of the Region Haná for Biotechnological and Agricultural Research, CZ 78371, Olomouc, Czech Republic.
authors.forename: Eva
authors.initials: E
authors.lastname: Hribova
authors.affiliation: Department of Plant Pathology, Wheat Genetics Resource Center, Kansas State University Manhattan, Manhattan, KS, 66506, USA.
authors.forename: Nidhi
authors.initials: N
authors.lastname: Rawat
authors.affiliation: Department of Plant Pathology, Wheat Genetics Resource Center, Kansas State University Manhattan, Manhattan, KS, 66506, USA.
authors.forename: Bhanu
authors.initials: B
authors.lastname: Kalia
authors.affiliation: Department of Plant Pathology, Wheat Genetics Resource Center, Kansas State University Manhattan, Manhattan, KS, 66506, USA.
authors.forename: Narinder
authors.initials: N
authors.lastname: Singh
authors.affiliation: Department of Plant Pathology, Wheat Genetics Resource Center, Kansas State University Manhattan, Manhattan, KS, 66506, USA.
authors.forename: Bernd
authors.initials: B
authors.lastname: Friebe
authors.affiliation: Institute of Experimental Botany, Centre of the Region Haná for Biotechnological and Agricultural Research, CZ 78371, Olomouc, Czech Republic.
authors.forename: Jaroslav
authors.initials: J
authors.lastname: Doležel
authors.affiliation: Department of Plant Pathology, Kansas State University, Manhattan, KS, 66502, USA.
authors.forename: Eduard
authors.initials: E
authors.lastname: Akhunov
authors.affiliation: Department of Plant Pathology, Wheat Genetics Resource Center, Kansas State University Manhattan, Manhattan, KS, 66506, USA.
authors.forename: Jesse
authors.initials: J
authors.lastname: Poland
authors.affiliation: Biotechnology Research Group, Department of Biological Sciences, Faculty of Science, King Abdulaziz University, Jeddah, 21589, Saudi Arabia.
authors.forename: Jamal S M
authors.initials: JS
authors.lastname: Sabir
authors.affiliation: Department of Plant Pathology, Wheat Genetics Resource Center, Kansas State University Manhattan, Manhattan, KS, 66506, USA.
authors.forename: Bikram S
authors.initials: BS
authors.lastname: Gill
mesh_terms: D058431:Brachypodium / Q000235:genetics; D002874:Chromosome Mapping; D032461:Chromosomes, Plant / Q000235:genetics*; D019143:Evolution, Molecular; D023061:Gene Order; D017343:Genes, Plant / Q000235:genetics*; D018745:Genome, Plant / Q000235:genetics*; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods; D001467:Hordeum / Q000235:genetics; D017404:In Situ Hybridization, Fluorescence; D012275:Oryza / Q000235:genetics; D006109:Poaceae / Q000145:classification / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide; D045868:Sorghum / Q000235:genetics; D014908:Triticum / Q000235:genetics
keywords: GenomeZipper; SNPs; flow sorting; next generation sequencing; single gene FISH; synteny