Rice, one of the world's most important food plants, has important syntenic relationships with the other cereal species and is a model plant for the grasses. Here we present a map-based, finished quality sequence that covers 95% of the 389 Mb genome, including virtually all of the euchromatin and two complete centromeres. A total of 37,544 nontransposable-element-related protein-coding genes were identified, of which 71% had a putative homologue in Arabidopsis. In a reciprocal analysis, 90% of the Arabidopsis proteins had a putative homologue in the predicted rice proteome. Twenty-nine per cent of the 37,544 predicted genes appear in clustered gene families. The number and classes of transposable elements found in the rice genome are consistent with the expansion of syntenic regions in the maize and sorghum genomes. We find evidence for widespread and recurrent gene transfer from the organelles to the nuclear chromosomes. The map-based sequence has proven useful for the identification of genes underlying agronomic traits. The additional single-nucleotide polymorphisms and simple sequence repeats identified in our study should accelerate improvements in rice production.

These authors contributed equally to this work. SummaryWe constructed physical maps of rice chromosomes 1, 2, and 6±9 with P1-derived arti®cial chromosome (PAC) and bacterial arti®cial chromosome (BAC) clones. These maps, with only 20 gaps, cover more than 97% of the predicted length of the six chromosomes. We submitted a total of 193 Mbp of non-overlapping sequences to public databases. We analyzed the DNA sequences of 1316 genetic markers and six centromere-speci®c repeats to facilitate characterization of chromosomal recombination frequency and of the genomic composition and structure of the centromeric regions. We found marked changes in the relative recombination rate along the length of each chromosome. Chromosomal recombination at the centromere core and surrounding regions on the six chromosomes was completely suppressed. These regions have a total physical length of about 23 Mbp, corresponding to 11.4% of the entire size of the six chromosomes. Chromosome 6 has the longest quiescent region, with about 5.6 Mbp, followed by chromosome 8, with quiescent region about half this size. Repetitive sequences accounted for at least 40% of the total genomic sequence on the partly sequenced centromeric region of chromosome 1. Rice CentO satellite DNA is arrayed in clusters and is closely associated with the presence of Centromeric Retrotransposon of Rice (CRR )-and RIce RetroElement 7 (RIRE7 )-like retroelement sequences. We also detected relatively small coldspot regions outside the centromeric region; their repetitive content and gene density were similar to those of regions with normal recombination rates. Sequence analysis of these regions suggests that either the amount or the organization patterns of repetitive sequences may play a role in the inactivation of recombination.

Understanding the organization of eukaryotic centromeres has both fundamental and applied importance because of their roles in chromosome segregation, karyotypic stability, and artificial chromosome-based cloning and expression vectors. Using clone-by-clone sequencing methodology, we obtained the complete genomic sequence of the centromeric region of rice (Oryza sativa) chromosome 8. Analysis of 1.97 Mb of contiguous nucleotide sequence revealed three large clusters of CentO satellite repeats (68.5 kb of 155-bp repeats) and >220 transposable element (TE)-related sequences; together, these account for~60% of this centromeric region. The 155-bp repeats were tandemly arrayed head to tail within the clusters, which had different orientations and were interrupted by TE-related sequences. The individual 155-bp CentO satellite repeats showed frequent transitions and transversions at eight nucleotide positions. The 40 TE elements with highly conserved sequences were mostly gypsy-type retrotransposons. Furthermore, 48 genes, showing high BLAST homology to known proteins or to rice full-length cDNAs, were predicted within the region; some were close to the CentO clusters. We then performed a genome-wide survey of the sequences and organization of CentO and RIRE7 families. Our study provides the complete sequence of a centromeric region from either plants or animals and likely will provide insight into the evolutionary and functional analysis of plant centromeres.

We present here the annotation of the complete genome of rice Oryza sativa L. ssp. japonica cultivar Nipponbare. All functional annotations for proteins and non-protein-coding RNA (npRNA) candidates were manually curated. Functions were identified or inferred in 19,969 (70%) of the proteins, and 131 possible npRNAs (including 58 antisense transcripts) were found. Almost 5000 annotated protein-coding genes were found to be disrupted in insertional mutant lines, which will accelerate future experimental validation of the annotations. The rice loci were determined by using cDNA sequences obtained from rice and other representative cereals. Our conservative estimate based on these loci and an extrapolation suggested that the gene number of rice is ∼32,000, which is smaller than previous estimates. We conducted comparative analyses between rice and Arabidopsis thaliana and found that both genomes possessed several lineage-specific genes, which might account for the observed differences between these species, while they had similar sets of predicted functional domains among the protein sequences. A system to control translational efficiency seems to be conserved across large evolutionary distances. Moreover, the evolutionary process of protein-coding genes was examined. Our results suggest that natural selection may have played a role for duplicated genes in both species, so that duplication was suppressed or favored in a manner that depended on the function of a gene.

Active DNA transposons are important tools for gene functional analysis. The endogenous non-autonomous transposon, nDart1-0, in rice (Oryza sativa L.) is expected to generate various transposon-insertion mutants because nDart1-0 elements tend to insert into genic regions under natural growth conditions. We have developed a specific method (nDart1-0-iPCR) for efficient detection of nDart1-0 insertions and successfully identified the SNOW-WHITE LEAF1 (SWL1) gene in a variegated albino (swl1-v) mutant obtained from the nDart1-promoted rice tagging line. The variegated albino phenotype was caused by insertion and excision of nDart1-0 in the 5'-untranslated region of the SWL1 gene predicted to encode an unknown protein with the N-terminal chloroplast transit peptide. SWL1 expression was detected in various rice tissues at different developmental stages. However, immunoblot analysis indicated that SWL1 protein accumulation was strictly regulated in a tissue-specific manner. In the swl1 mutant, formations of grana and stroma thylakoids and prolamellar bodies were inhibited. This study revealed that SWL1 is essential for the beginning of thylakoid membrane organization during chloroplast development. Furthermore, we provide a developmental perspective on the nDart1-promoted tagging line to characterize unidentified gene functions in rice.

A non-autonomous DNA transposon in rice, nDart1, is actively transposed in the presence of an autonomous element, aDart1, under natural conditions. The nDart1-promoted gene tagging line was developed using the endogenous nDart1/aDart1 system to generate various rice mutants effectively. While the dominant mutants were occasionally isolated from the tagging line, it was unclear what causes dominant mutations. A semidominant mutant, Bushy dwarf tiller1 (Bdt1), which has the valuable agronomic traits of multiple tillering and dwarfism, was obtained from the tagging line. Bdt1 mutant carried a newly inserted nDart1 at 38-bp upstream of transcription initiation site of a non-protein-coding gene, miR156d. This insertion caused an upstream shift of the miR156d transcription initiation site and, consequently, increased the functional transcripts producing mature microRNAs. These results indicate that the total amount of miR156d is controlled not only by transcript quantity but also by transcript quality. Furthermore, transgenic lines introduced an miR156d fragment that flanked the nDart1 sequence at the 5′ region, suggesting that insertion of nDart1 in the gene promoter region enhances gene expression as a cis-element. This study demonstrates the ability of nDart1 to produce gain-of-function mutants as well as further insights into the function of transposable elements in genome evolution.

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