William Nelson

Genome sequence of the palaeopolyploid soybean

Soybean (Glycine max) is one of the most important crop plants for seed protein and oil content, and for its capacity to fix atmospheric nitrogen through symbioses with soil-borne microorganisms. We sequenced the 1.1-gigabase genome by a whole-genome shotgun approach and integrated it with physical and high-density genetic maps to create a chromosome-scale draft sequence assembly. We predict 46,430 protein-coding genes, 70% more than Arabidopsis and similar to the poplar genome which, like soybean, is an ancient polyploid (palaeopolyploid). About 78% of the predicted genes occur in chromosome ends, which comprise less than one-half of the genome but account for nearly all of the genetic recombination. Genome duplications occurred at approximately 59 and 13 million years ago, resulting in a highly duplicated genome with nearly 75% of the genes present in multiple copies. The two duplication events were followed by gene diversification and loss, and numerous chromosome rearrangements. An accurate soybean genome sequence will facilitate the identification of the genetic basis of many soybean traits, and accelerate the creation of improved soybean varieties.

Physical and genetic structure of the maize genome reflects its complex evolutionary history.

Maize (Zea mays L.) is one of the most important cereal crops and a model for the study of genetics, evolution, and domestication. To better understand maize genome organization and to build a framework for genome sequencing, we constructed a sequence-ready fingerprinted contig-based physical map that covers 93.5% of the genome, of which 86.1% is aligned to the genetic map. The fingerprinted contig map contains 25,908 genic markers that enabled us to align nearly 73% of the anchored maize genome to the rice genome. The distribution pattern of expressed sequence tags correlates to that of recombination. In collinear regions, 1 kb in rice corresponds to an average of 3.2 kb in maize, yet maize has a 6-fold genome size expansion. This can be explained by the fact that most rice regions correspond to two regions in maize as a result of its recent polyploid origin. Inversions account for the majority of chromosome structural variations during subsequent maize diploidization. We also find clear evidence of ancient genome duplication predating the divergence of the progenitors of maize and rice. Reconstructing the paleoethnobotany of the maize genome indicates that the progenitors of modern maize contained ten chromosomes.

SyMAP v3.4: a turnkey synteny system with application to plant genomes

SyMAP (Synteny Mapping and Analysis Program) was originally developed to compute synteny blocks between a sequenced genome and a FPC map, and has been extended to support pairs of sequenced genomes. SyMAP uses MUMmer to compute the raw hits between the two genomes, which are then clustered and filtered using the optional gene annotation. The filtered hits are input to the synteny algorithm, which was designed to discover duplicated regions and form larger-scale synteny blocks, where intervening micro-rearrangements are allowed. SyMAP provides extensive interactive Java displays at all levels of resolution along with simultaneous displays of multiple aligned pairs. The synteny blocks from multiple chromosomes may be displayed in a high-level dot plot or three-dimensional view, and the user may then drill down to see the details of a region, including the alignments of the hits to the gene annotation. These capabilities are illustrated by showing their application to the study of genome duplication, differential gene loss and transitive homology between sorghum, maize and rice. The software may be used from a website or standalone for the best performance. A project manager is provided to organize and automate the analysis of multi-genome groups. The software is freely distributed at http://www.agcol.arizona.edu/software/symap.

Whole-genome validation of high-information-content fingerprinting

Fluorescent-based high-information-content fingerprinting (HICF) techniques have recently been developed for physical mapping. These techniques make use of automated capillary DNA sequencing instruments to enable both high-resolution and high-throughput fingerprinting. In this article, we report the construction of a whole-genome HICF FPC map for maize (Zea mays subsp. mays cv B73), using a variant of HICF in which a type IIS restriction enzyme is used to generate the fluorescently labeled fragments. The HICF maize map was constructed from the same three maize bacterial artificial chromosome libraries as previously used for the whole-genome agarose FPC map, providing a unique opportunity for direct comparison of the agarose and HICF methods; as a result, it was found that HICF has substantially greater sensitivity in forming contigs. An improved assembly procedure is also described that uses automatic end-merging of contigs to reduce the effects of contamination and repetitive bands. Several new features in FPC v7.2 are presented, including shared-memory multiprocessing, which allows dramatically faster assemblies, and automatic end-merging, which permits more accurate assemblies. It is further shown that sequenced clones may be digested in silico and located accurately on the HICF assembly, despite size deviations that prevent the precise prediction of experimental fingerprints. Finally, repetitive bands are isolated, and their effect on the assembly is studied.

Life at the hyperarid margin: novel bacterial diversity in arid soils of the Atacama Desert, Chile

Nearly half the earth’s surface is occupied by dryland ecosystems, regions susceptible to reduced states of biological productivity caused by climate fluctuations. Of these regions, arid zones located at the interface between vegetated semiarid regions and biologically unproductive hyperarid zones are considered most vulnerable. The objective of this study was to conduct a deep diversity analysis of bacterial communities in unvegetated arid soils of the Atacama Desert, to characterize community structure and infer the functional potential of these communities based on observed phylogenetic associations. A 454-pyrotag analysis was conducted of three unvegetated arid sites located at the hyperarid–arid margin. The analysis revealed communities with unique bacterial diversity marked by high abundances of novel Actinobacteria and Chloroflexi and low levels of Acidobacteria and Proteobacteria, phyla that are dominant in many biomes. A 16S rRNA gene library of one site revealed the presence of clones with phylogenetic associations to chemoautotrophic taxa able to obtain energy through oxidation of nitrite, carbon monoxide, iron, or sulfur. Thus, soils at the hyperarid margin were found to harbor a wealth of novel bacteria and to support potentially viable communities with phylogenetic associations to non-phototrophic primary producers and bacteria capable of biogeochemical cycling.

Construction, alignment and analysis of twelve framework physical maps that represent the ten genome types of the genus Oryza

We describe the establishment and analysis of a genus-wide comparative framework composed of 12 bacterial artificial chromosome fingerprint and end-sequenced physical maps representing the 10 genome types of Oryza aligned to the O. sativa ssp. japonica reference genome sequence. Over 932 Mb of end sequence was analyzed for repeats, simple sequence repeats, miRNA and single nucleotide variations, providing the most extensive analysis of Oryza sequence to date.

Making a living while starving in the dark: metagenomic insights into the energy dynamics of a carbonate cave.

Carbonate caves represent subterranean ecosystems that are largely devoid of phototrophic primary production. In semiarid and arid regions, allochthonous organic carbon inputs entering caves with vadose-zone drip water are minimal, creating highly oligotrophic conditions; however, past research indicates that carbonate speleothem surfaces in these caves support diverse, predominantly heterotrophic prokaryotic communities. The current study applied a metagenomic approach to elucidate the community structure and potential energy dynamics of microbial communities, colonizing speleothem surfaces in Kartchner Caverns, a carbonate cave in semiarid, southeastern Arizona, USA. Manual inspection of a speleothem metagenome revealed a community genetically adapted to low-nutrient conditions with indications that a nitrogen-based primary production strategy is probable, including contributions from both Archaea and Bacteria. Genes for all six known CO2-fixation pathways were detected in the metagenome and RuBisCo genes representative of the Calvin–Benson–Bassham cycle were over-represented in Kartchner speleothem metagenomes relative to bulk soil, rhizosphere soil and deep-ocean communities. Intriguingly, quantitative PCR found Archaea to be significantly more abundant in the cave communities than in soils above the cave. MEtaGenome ANalyzer (MEGAN) analysis of speleothem metagenome sequence reads found Thaumarchaeota to be the third most abundant phylum in the community, and identified taxonomic associations to this phylum for indicator genes representative of multiple CO2-fixation pathways. The results revealed that this oligotrophic subterranean environment supports a unique chemoautotrophic microbial community with potentially novel nutrient cycling strategies. These strategies may provide key insights into other ecosystems dominated by oligotrophy, including aphotic subsurface soils or aquifers and photic systems such as arid deserts.

A BAC-based physical map of Brachypodium distachyon and its comparative analysis with rice and wheat

BackgroundBrachypodium distachyon (Brachypodium) has been recognized as a new model species for comparative and functional genomics of cereal and bioenergy crops because it possesses many biological attributes desirable in a model, such as a small genome size, short stature, self-pollinating habit, and short generation cycle. To maximize the utility of Brachypodiu m as a model for basic and applied research it is necessary to develop genomic resources for it. A BAC-based physical map is one of them. A physical map will facilitate analysis of genome structure, comparative genomics, and assembly of the entire genome sequence.ResultsA total of 67,151 Brachypodium BAC clones were fingerprinted with the SNaPshot HICF fingerprinting method and a genome-wide physical map of the Brachypodium genome was constructed. The map consisted of 671 contigs and 2,161 clones remained as singletons. The contigs and singletons spanned 414 Mb. A total of 13,970 gene-related sequences were detected in the BAC end sequences (BES). These gene tags aligned 345 contigs with 336 Mb of rice genome sequence, showing that Brachypodium and rice genomes are generally highly colinear. Divergent regions were mainly in the rice centromeric regions. A dot-plot of Brachypodium contigs against the rice genome sequences revealed remnants of the whole-genome duplication caused by paleotetraploidy, which were previously found in rice and sorghum. Brachypodium contigs were anchored to the wheat deletion bin maps with the BES gene-tags, opening the door to Brachypodium-Triticeae comparative genomics.ConclusionThe construction of the Brachypodium physical map, and its comparison with the rice genome sequence demonstrated the utility of the SNaPshot-HICF method in the construction of BAC-based physical maps. The map represents an important genomic resource for the completion of Brachypodium genome sequence and grass comparative genomics. A draft of the physical map and its comparisons with rice and wheat are available at http://phymap.ucdavis.edu/brachypodium/.

Comparative Physical Mapping Between Oryza sativa (AA Genome Type) and O. punctata (BB Genome Type)

A comparative physical map of the AA genome (Oryza sativa) and the BB genome (O. punctata) was constructed by aligning a physical map of O. punctata, deduced from 63,942 BAC end sequences (BESs) and 34,224 fingerprints, onto the O. sativa genome sequence. The level of conservation of each chromosome between the two species was determined by calculating a ratio of BES alignments. The alignment result suggests more divergence of intergenic and repeat regions in comparison to gene-rich regions. Further, this characteristic enabled localization of heterochromatic and euchromatic regions for each chromosome of both species. The alignment identified 16 locations containing expansions, contractions, inversions, and transpositions. By aligning 40% of the punctata BES on the map, 87% of the punctata FPC map covered 98% of the O. sativa genome sequence. The genome size of O. punctata was estimated to be 8% larger than that of O. sativa with individual chromosome differences of 1.5–16.5%. The sum of expansions and contractions observed in regions >500 kb were similar, suggesting that most of the contractions/expansions contributing to the genome size difference between the two species are small, thus preserving the macro-collinearity between these species, which diverged ∼2 million years ago.

Integrating sequence with FPC fingerprint maps

Recent advances in both clone fingerprinting and draft sequencing technology have made it increasingly common for species to have a bacterial artificial clone (BAC) fingerprint map, BAC end sequences (BESs) and draft genomic sequence. The FPC (fingerprinted contigs) software package contains three modules that maximize the value of these resources. The BSS (blast some sequence) module provides a way to easily view the results of aligning draft sequence to the BESs, and integrates the results with the following two modules. The MTP (minimal tiling path) module uses sequence and fingerprints to determine a minimal tiling path of clones. The DSI (draft sequence integration) module aligns draft sequences to FPC contigs, displays them alongside the contigs and identifies potential discrepancies; the alignment can be based on either individual BES alignments to the draft, or on the locations of BESs that have been assembled into the draft. FPC also supports high-throughput fingerprint map generation as its time-intensive functions have been parallelized for Unix-based desktops or servers with multiple CPUs. Simulation results are provided for the MTP, DSI and parallelization. These features are in the FPC V9.3 software package, which is freely available.