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Next-generation sequencing and syntenic integration of flow-sorted arms of wheat chromosome 4A exposes the chromosome structure and gene content
Pilar Hernandez, Mihaela Martis, Gabriel Dorado, Matthias Pfeifer, Sergio Gálvez, Sebastian Schaaf, Nicolás Jouve, Hana Šimková, Miroslav Valárik, Jaroslav Doležel, Klaus F.X. Mayer
Wheat is the third most important crop for human nutrition in the world. The availability of high-resolution genetic and physical maps and ultimately a complete genome sequence holds great promise for breeding improved varieties to cope with increasing food demand under the conditions of changing global climate. However, the large size of the bread wheat (Triticum aestivum) genome (approximately 17 Gb/1C) and the triplication of genic sequence resulting from its hexaploid status have impeded genome sequencing of this important crop species. Here we describe the use of mitotic chromosome flow sorting to separately purify and then shotgun-sequence a pair of telocentric chromosomes that together form chromosome 4A (856 Mb/1C) of wheat. The isolation of this much reduced template and the consequent avoidance of the problem of sequence duplication, in conjunction with synteny-based comparisons with other grass genomes, have facilitated construction of an ordered gene map of chromosome 4A, embracing ≥85% of its total gene content, and have enabled precise localization of the various translocation and inversion breakpoints on chromosome 4A that differentiate it from its progenitor chromosome in the A genome diploid donor. The gene map of chromosome 4A, together with the emerging sequences of homoeologous wheat chromosome groups 4, 5 and 7, represent unique resources that will allow us to obtain new insights into the evolutionary dynamics between homoeologous chromosomes and syntenic chromosomal regions.
Sequence Composition and Gene Content of the Short
Arm of Rye (Secale cereale) Chromosome 1
Silvia Fluch, Dieter Kopecky, Kornel Burg, Hana Šimková, Stefan Taudien, Andreas Petzold, Marie Kubaláková, Matthias Platzer, Maria Berenyi, Siegfried Krainer, Jaroslav Doležel, Tamas Lelley
Background: The purpose of the study is to elucidate the sequence composition of the short arm of rye chromosome 1(Secale cereale) with special focus on its gene content, because this portion of the rye genome is an integrated part of several hundreds of bread wheat varieties worldwide.
Methodology/Principal Findings: Multiple Displacement Amplification of 1RS DNA, obtained from flow sorted 1RS chromosomes, using 1RS ditelosomic wheat-rye addition line, and subsequent Roche 454FLX sequencing of this DNA yielded 195,313,589 bp sequence information. This quantity of sequence information resulted in 0.436sequence coverage of the 1RS chromosome arm, permitting the identification of genes with estimated probability of 95%. A detailed analysis revealed that more than 5% of the 1RS sequence consisted of gene space, identifying at least 3,121 gene loci representing
1,882 different gene functions. Repetitive elements comprised about 72% of the 1RS sequence, Gypsy/Sabrina (13.3%) being the most abundant. More than four thousand simple sequence repeat (SSR) sites mostly located in gene related sequence reads were identified for possible marker development. The existence of chloroplast insertions in 1RS has been verified by identifying chimeric chloroplast-genomic sequence reads. Synteny analysis of 1RS to the full genomes of Oryza sativa and Brachypodium distachyon revealed that about half of the genes of 1RS correspond to the distal end of the short
arm of rice chromosome 5 and the proximal region of the long arm of Brachypodium distachyon chromosome 2. Comparison of the gene content of 1RS to 1HS barley chromosome arm revealed high conservation of genes related to chromosome 5 of rice.
Conclusions: The present study revealed the gene content and potential gene functions on this chromosome arm and demonstrated numerous sequence elements like SSRs and gene-related sequences, which can be utilised for future research as well as in breeding of wheat and rye.
A 3,000-Loci Transcription Map of Chromosome 3B
Unravels the Structural and Functional Features of Gene
Islands in Hexaploid Wheat
Camille Rustenholz, Frédéric Choulet, Christel Laugier, Jan Šafář, Hana Šimková, Jaroslav Doležel, Federica Magni, Simone Scalabrin, Federica Cattonaro, Sonia Vautrin, Arnaud Bellec, Hélène Bergés, Catherine Feuillet, and Etienne Paux
To improve our understanding of the organization and regulation of the wheat (Triticum aestivum) gene space, we established a transcription map of a wheat chromosome (3B) by hybridizing a newly developed wheat expression microarray with bacterial artificial chromosome pools from a new version of the 3B physical map as well as with cDNA probes derived from 15 RNA samples. Mapping data for almost 3,000 genes showed that the gene space spans the whole chromosome 3B with a 2-fold increase of gene density toward the telomeres due to an increase in the number of genes in islands. Comparative analyses with rice (Oryza sativa) and Brachypodium distachyon revealed that these gene islands are composed mainly of genes likely originating from interchromosomal gene duplications. Gene Ontology and expression profile analyses for the 3,000 genes located along the chromosome revealed that the gene islands are enriched significantly in genes sharing the same function or expression profile, thereby suggesting that genes in islands acquired shared regulation during evolution. Only a small fraction of these clusters of cofunctional and coexpressed genes was conserved with rice and B. distachyon, indicating a recent origin. Finally, genes with the same expression profiles in remote islands (coregulation islands) were identified suggesting long-distance regulation of gene expression along the chromosomes in wheat.
