Effective Isolation of Retrotransposons and Repetitive DNA Families from the Wheat Genome

New classes of repetitive DNA elements were effectively identified by isolating small fragments of the elements from the wheat genome. A wheat A genome library was constructed from Triticum monococcum by degenerate cleavage with EcoO1091, the recognition sites of which consisted of 5＇-PuGGNCCPy-3＇ multi-sequences. Three novel repetitive sequences pTm6, pTm69 and pTm58 derived from the A genome were screened and tested for high copy number using a blotting approach, pTm6 showed identity with integrase domains of the barley Tyl-Copia-retrotransposon BARE-1 and pTm58 showed similarity to the barley Ty3-gypsy-like retrotransposon Romani. pTm69, however, constituted a tandem array with useful genomic specificities, but did not share any identity with known repetitive elements. This study also sought to isolate wheat D-genome-specific repetitive elements regardless of the level of methylation, by genomic subtraction. Total genomic DNA of Aegilops tauschii was cleaved into short fragments with a methylation-insensitive 4 bp cutter, Mbol, and then common DNA sequences between Ae. tauschii and Triticum turgidum were subtracted by annealing with excess T. turgidum genomic DNA. The D genome repetitive sequence pAt1 was isolated and used to identify an additional novel repetitive sequence family from wheat bacterial artificial chromosomes with a size range of 1 395-1 850 bp. The methods successfully led pathfinding of two unique repetitive families.

Integrating the physical and genetic map of bread wheat facilitates the detection of chromosomal rearrangements

The bread wheat genome harbors a high content of repetitive DNA,which is amenable to detection and characterization using fluorescence in situ hybridization(FISH)karyotyping.An integrated genetic map was derived from a recombinant inbred population bred from a cross between a synthetic hexaploid wheat and a commercial Chinese bread wheat cultivar,based on 28 variable FISH sites and>150000 single nucleotide polymorphism(SNP)loci.The majority(20/28)of the variable FISH sites were physically located within a chromosomal region consistent with the genetic location inferred from that of their co-segregating SNP loci.The eight exceptions reflected the presence of either a translocation(1 R/1 B,1 A/7 A)or a presumptive intra-chromosomal inversion(4 A).For eight out of the nine FISH sites detected on the Chinese Spring(CS)karyotype,there was a good match with the reference genome sequence,indicating that the most recent assembly has dealt well with the problem of placing tandem repeats.The integrated genetic map produced for wheat is informative as to the location of blocks of tandemly repeated DNA and can aid in improving the quality of the genome sequence assembly in regions surrounding these blocks.

Deciphering the evolution and complexity of wheat germplasm from a genomic perspective

Bread wheat provides an essential fraction of the daily calorific intake for humanity.Due to its huge and complex genome,progress in studying on the wheat genome is substantially trailed behind those of the other two major crops,rice and maize,for at least a decade.With rapid advances in genome assembling and reduced cost of high-throughput sequencing,emerging de novo genome assemblies of wheat and whole-genome sequencing data are leading to a paradigm shift in wheat research.Here,we review recent progress in dissecting the complex genome and germplasm evolution of wheat since the release of the first high-quality wheat genome.New insights have been gained in the evolution of wheat germplasm during domestication and modern breeding progress,genomic variations at multiple scales contributing to the diversity of wheat germplasm,and complex transcriptional and epigenetic regulations of functional genes in polyploid wheat.Genomics databases and bioinformatics tools meeting the urgent needs of wheat ge-nomics research are also summarized.The ever-increasing omics data,along with advanced tools and well-structured databases,are expected to accelerate deciphering the germplasm and gene resources in wheat for future breeding advances.