Simone L. Macmil

Sequencing of Medicago truncatula, a model organism of legume biology, shows that genome duplications had a role in the evolution of endosymbiotic nitrogen fixation. Legumes are unusual among plants in that they can carry out endosymbiotic nitrogen fixation with rhizobial bacteria. The genome of Medicago truncatula (also known as barrel medic or barrel clover), a well-established model for the study of legume biology, has now been sequenced. Genome analysis shows that M. truncatula has undergone several rounds of whole-genome duplication, and that the duplication that took place approximately 58 million years ago played an important part in the evolution of endosymbiotic nitrogen fixation. Legumes (Fabaceae or Leguminosae) are unique among cultivated plants for their ability to carry out endosymbiotic nitrogen fixation with rhizobial bacteria, a process that takes place in a specialized structure known as the nodule. Legumes belong to one of the two main groups of eurosids, the Fabidae, which includes most species capable of endosymbiotic nitrogen fixation1. Legumes comprise several evolutionary lineages derived from a common ancestor 60 million years ago (Myr ago). Papilionoids are the largest clade, dating nearly to the origin of legumes and containing most cultivated species2. Medicago truncatula is a long-established model for the study of legume biology. Here we describe the draft sequence of the M. truncatula euchromatin based on a recently completed BAC assembly supplemented with Illumina shotgun sequence, together capturing ∼94% of all M. truncatula genes. A whole-genome duplication (WGD) approximately 58 Myr ago had a major role in shaping the M. truncatula genome and thereby contributed to the evolution of endosymbiotic nitrogen fixation. Subsequent to the WGD, the M. truncatula genome experienced higher levels of rearrangement than two other sequenced legumes, Glycine max and Lotus japonicus. M. truncatula is a close relative of alfalfa (Medicago sativa), a widely cultivated crop with limited genomics tools and complex autotetraploid genetics. As such, the M. truncatula genome sequence provides significant opportunities to expand alfalfa’s genomic toolbox.

Herbivorous insects use detoxification enzymes, including cytochrome P450 monooxygenases, glutathione S-transferases, and carboxy/cholinesterases, to metabolize otherwise deleterious plant secondary metabolites. Whereas Acyrthosiphon pisum (pea aphid) feeds almost exclusively from the Fabaceae, Myzus persicae (green peach aphid) feeds from hundreds of species in more than forty plant families. Therefore, M. persicae as a species would be exposed to a greater diversity of plant secondary metabolites than A. pisum, and has been predicted to require a larger complement of detoxification enzymes. A comparison of M. persicae cDNA and A. pisum genomic sequences is partially consistent with this hypothesis. There is evidence of at least 40% more cytochrome P450 genes in M. persicae than in A. pisum. In contrast, no major differences were found between the two species in the numbers of glutathione S-transferases, and carboxy/cholinesterases. However, given the incomplete M. persicae cDNA data set, the number of identified detoxification genes in this species is likely to be an underestimate.

BACKGROUND: The draft genome sequence of the ascidian Ciona intestinalis, along with associated gene models, has been a valuable research resource. However, recently accumulated expressed sequence tag (EST)/cDNA data have revealed numerous inconsistencies with the gene models due in part to intrinsic limitations in gene prediction programs and in part to the fragmented nature of the assembly. RESULTS: We have prepared a less-fragmented assembly on the basis of scaffold-joining guided by paired-end EST and bacterial artificial chromosome (BAC) sequences, and BAC chromosomal in situ hybridization data. The new assembly (115.2 Mb) is similar in length to the initial assembly (116.7 Mb) but contains 1,272 (approximately 50%) fewer scaffolds. The largest scaffold in the new assembly incorporates 95 initial-assembly scaffolds. In conjunction with the new assembly, we have prepared a greatly improved global gene model set strictly correlated with the extensive currently available EST data. The total gene number (15,254) is similar to that of the initial set (15,582), but the new set includes 3,330 models at genomic sites where none were present in the initial set, and 1,779 models that represent fusions of multiple previously incomplete models. In approximately half, 5'-ends were precisely mapped using 5'-full-length ESTs, an important refinement even in otherwise unchanged models. CONCLUSION: Using these new resources, we identify a population of non-canonical (non-GT-AG) introns and also find that approximately 20% of Ciona genes reside in operons and that operons contain a high proportion of single-exon genes. Thus, the present dataset provides an opportunity to analyze the Ciona genome much more precisely than ever.

BACKGROUND: Sugarcane (Saccharum spp.) has become an increasingly important crop for its leading role in biofuel production. The high sugar content species S. officinarum is an octoploid without known diploid or tetraploid progenitors. Commercial sugarcane cultivars are hybrids between S. officinarum and wild species S. spontaneum with ploidy at approximately 12x. The complex autopolyploid sugarcane genome has not been characterized at the DNA sequence level. RESULTS: The microsynteny between sugarcane and sorghum was assessed by comparing 454 pyrosequences of 20 sugarcane bacterial artificial chromosomes (BACs) with sorghum sequences. These 20 BACs were selected by hybridization of 1961 single copy sorghum overgo probes to the sugarcane BAC library with one sugarcane BAC corresponding to each of the 20 sorghum chromosome arms. The genic regions of the sugarcane BACs shared an average of 95.2% sequence identity with sorghum, and the sorghum genome was used as a template to order sequence contigs covering 78.2% of the 20 BAC sequences. About 53.1% of the sugarcane BAC sequences are aligned with sorghum sequence. The unaligned regions contain non-coding and repetitive sequences. Within the aligned sequences, 209 genes were annotated in sugarcane and 202 in sorghum. Seventeen genes appeared to be sugarcane-specific and all validated by sugarcane ESTs, while 12 appeared sorghum-specific but only one validated by sorghum ESTs. Twelve of the 17 sugarcane-specific genes have no match in the non-redundant protein database in GenBank, perhaps encoding proteins for sugarcane-specific processes. The sorghum orthologous regions appeared to have expanded relative to sugarcane, mostly by the increase of retrotransposons. CONCLUSIONS: The sugarcane and sorghum genomes are mostly collinear in the genic regions, and the sorghum genome can be used as a template for assembling much of the genic DNA of the autopolyploid sugarcane genome. The comparable gene density between sugarcane BACs and corresponding sorghum sequences defied the notion that polyploidy species might have faster pace of gene loss due to the redundancy of multiple alleles at each locus.

MicroRNAs (miRNAs) and small-interfering RNAs (siRNAs) have emerged as important regulators of gene expression in higher eukaryotes. Recent studies indicate that genomes in higher plants encode lineage-specific and species-specific miRNAs in addition to the well-conserved miRNAs. Leguminous plants are grown throughout the world for food and forage production. To date the lack of genomic sequence data has prevented systematic examination of small RNAs in leguminous plants. Medicago truncatula, a diploid plant with a near-completely sequenced genome has recently emerged as an important model legume. We sequenced a small RNA library generated from M. truncatula to identify not only conserved miRNAs but also novel small RNAs, if any. Eight novel small RNAs were identified, of which four (miR1507, miR2118, miR2119 and miR2199) are annotated as legume-specific miRNAs because these are conserved in related legumes. Three novel transcripts encoding TIR-NBS-LRR proteins are validated as targets for one of the novel miRNA, miR2118. Small RNA sequence analysis coupled with the small RNA blot analysis, confirmed the expression of around 20 conserved miRNA families in M. truncatula. Fifteen transcripts have been validated as targets for conserved miRNAs. We also characterized Tas3-siRNA biogenesis in M. truncatula and validated three auxin response factor (ARF) transcripts that are targeted by tasiRNAs. These findings indicate that M. truncatula and possibly other related legumes have complex mechanisms of gene regulation involving specific and common small RNAs operating post-transcriptionally.