稻属不同染色体组着丝粒BAC克隆的分离和鉴定

着丝粒在真核生物有丝分裂和减数分裂染色体正常的分离和传递中起着重要的作用。通过构建5个稻属二倍体野生种的基因组BAC文库, 采用菌落杂交和FISH技术, 筛选和鉴定了各染色体组着丝粒克隆, 并且分析了这些克隆在不同基因组间的共杂交情况, 结果表明: (1) C染色体组的野生种O. officinalis 和F染色体组的野生种O. brachyantha具有各自着丝粒特异的卫星DNA序列, 并且O. brachyantha着丝粒还具有特异的逆转座子序列; (2) A、B和E染色体组的野生稻O. glaberrima、O. punctata和O. australiensis着丝粒区域都含有与栽培稻着丝粒重复序列CentO和CRR同源的序列; (3) C染色体组野生稻O. officinalis的2条体细胞染色体着丝粒具有CentO的同源序列, 同时也发现其所有着丝粒区域都包含栽培稻CRR的同源序列。这些结果对克隆稻属不同染色体组的着丝粒序列、研究不同染色体组间着丝粒的进化关系和稻属不同着丝粒DNA序列与功能之间的关系均具有重要意义。     

Evolution of alu family repeats since the divergence of human and chimpanzee

Summary The DNA sequences of three members of the Alu family of repeated sequences located 5 to the chimpanzee 2 gene have been determined. The base sequences of the three corresponding human Alu family repeats have been previously determined, permitting the comparison of identical Alu family members in human and chimpanzee. Here we compare the sequences of seven pairs of chimpanzee and human Alu repeats. In each case, with the exception of minor sequence differences, the identical Alu repeat is located at identical sites in the human and chimpanzee genomes. The Alu repeats diverge at the rate expected for nonselected sequences. Sequence conversion has not replaced any of these 14 Alu family members since the divergence between chimpanzee and human.     

Diverse retrotransposon families and an AT-rich satellite DNA revealed in giant genomes of Fritillaria lilies

Background and Aims

The genus Fritillaria (Liliaceae) comprises species with extremely large genomes (1C = 30 000–127 000 Mb) and a bicontinental distribution. Most North American species (subgenus Liliorhiza) differ from Eurasian Fritillaria species by their distinct phylogenetic position and increased amounts of heterochromatin. This study examined the contribution of major repetitive elements to the genome obesity found in Fritillaria and identified repeats contributing to the heterochromatin arrays in Liliorhiza species.

Methods

Two Fritillaria species of similar genome size were selected for detailed analysis, one from each phylogeographical clade: F. affinis (1C = 45·6 pg, North America) and F. imperialis (1C = 43·0 pg, Eurasia). Fosmid libraries were constructed from their genomic DNAs and used for identification, sequence characterization, quantification and chromosome localization of clones containing highly repeated sequences.

Key Results and Conclusions

Repeats corresponding to 6·7 and 4·7 % of the F. affinis and F. imperialis genome, respectively, were identified. Chromoviruses and the Tat lineage of Ty3/gypsy group long terminal repeat retrotransposons were identified as the predominant components of the highly repeated fractions in the F. affinis and F. imperialis genomes, respectively. In addition, a heterogeneous, extremely AT-rich satellite repeat was isolated from F. affinis. The FriSAT1 repeat localized in heterochromatic bands makes up approx. 26 % of the F. affinis genome and substantial genomic fractions in several other Liliorhiza species. However, no evidence of a relationship between heterochromatin content and genome size variation was observed. Also, this study was unable to reveal any predominant repeats which tracked the increasing/decreasing trends of genome size evolution in Fritillaria. Instead, the giant Fritillaria genomes seem to be composed of many diversified families of transposable elements. We hypothesize that the genome obesity may be partly determined by the failure of removal mechanisms to counterbalance effectively the retrotransposon amplification.     

Constant and variable parts of the 155-bp centromeric repeat in Camptochironomus

Centromeres in dipteran insects belonging to the subgenus Camptochironomus (C. tentans and C. pallidivittatus) contain tandem repeats of a 155-bp repeat. A special structural feature of the 155-bp unit is a region with two palindromes connected by a short piece of DNA with only AT base pairs, which has at least a superficial similarity to a functionally important part of the Saccharomyces cerevisiae centromere. As a parameter for functional importance we have measured frequencies of mutations along the unit in samples of repeats from the two species. We find that it consists of about equal parts of highly conserved and considerably less-conserved DNA. The palindromes are localized in the conserved part of the repeat. Correspondence to: J.-E. Edström     

Abundance, polymorphism and genetic mapping of microsatellites in rice

Dinucleotide microsatellites have been characterized and used as genetic markers in rice. Screening of a rice genomic library with poly(dG-dA)·(dC-dT) and poly(dG-dT)·(dC-dA) probes indicated that (GA)_n repeats occurred, on average, once every 225 kb and (GT)_n repeats once every 480 kb. DNA sequencing of ten randomly selected microsatellites indicated that the numbers of repeats ranged from 12 to 34 and that the patterns of microsatellites in rice were similar to those of humans and other mammals. Primers to these microsatellite loci as well as to four published microsatellite-containing sequences have been designed and degrees of polymorphism has been examined with 20 rice accessions. Multiple alleles, ranging from 5 to 11, have been observed at all the microsatellite loci in 20 rice accessions. Alleles specific to two cultivated subspecies, indica and japonica, were found in some microsatellite loci. Heterozygosity values of all the microsatellite markers were significantly higher than those of RFLP markers, based upon a parallel comparison. Ten microsatellite loci have been genetically mapped to four rice chromosomes. The genomic distribution of microsatellites appears to be random in rice.     

Evolutionary dynamics of an ancient retrotransposon family provides insights into evolution of genome size in the genus Oryza

Long terminal repeat (LTR) retrotransposons constitute a significant portion of most eukaryote genomes and can dramatically change genome size and organization. Although LTR retrotransposon content variation is well documented, the dynamics of genomic flux caused by their activity are poorly understood on an evolutionary time scale. This is primarily because of the lack of an experimental system composed of closely related species whose divergence times are within the limits of the ability to detect ancestrally related retrotransposons. The genus Oryza, with 24 species, ten genome types, different ploidy levels and over threefold genome size variation, constitutes an ideal experimental system to explore genus-level transposon dynamics. Here we present data on the discovery and characterization of an LTR retrotransposon family named RWG in the genus Oryza. Comparative analysis of transposon content (approximately 20 to 27,000 copies) and transpositional history of this family across the genus revealed a broad spectrum of independent and lineage-specific changes that have implications for the evolution of genome size and organization. In particular, we provide evidence that the basal GG genome of Oryza (O. granulata) has expanded by nearly 25% by a burst of the RWG lineage Gran3 subsequent to speciation. Finally we describe the recent evolutionary origin of Dasheng, a large retrotransposon derivative of the RWG family, specifically found in the A, B and C genome lineages of Oryza.     

Structure of Kluyveromyces lactis subtelomeres: duplications and gene content

We have constructed a map of the duplicated regions of Kluyveromyces lactis subtelomeres. Seven out of 12 subtelomeres contain an almost identical 9 kb long segment starting from the end. This segment is bordered by a long terminal repeat element. Two of the subtelomeres share sequence similarity that extends over a total of 20 kb. The other subtelomeres also contain duplicated regions of 1-6 kb. Nonduplicated regions contain unique genes and genes from paralog families. All duplicated segments are in the same orientation with respect to the telomere, probably as a result of genetic exchange. We map the only two copies of retrotransposons in the genome, in subtelomeres. Low-complexity gene sequences that encode threonine- and serine-rich peptides are associated with the subtelomeres of K. lactis, as in Saccharomyces cerevisiae. The ubiquity of these sequences in hemiascomycete genomes, and the propensity they have to encode proteins with extracellular localization, make these genes ideal candidates for fast evolving 'contingency' genes involved in the adaptation of a species to its environment.     

Characterization of simple sequence repeat markers derived in silico from Brassica rapa bacterial artificial chromosome sequences and their application in Brassica napus

A collaborative Brassica rapa genome sequencing project is currently in progress to aid the identification of agronomically important traits in Brassica species. As an initial stage, the ends of over 110 000 bacterial artificial chromosome clones were sequenced and mined for simple sequence repeats (SSRs). We present the characterization of 40 of these SSRs and their application in Brassica napus. The markers were screened against six Brassica species and Arabidopsis, and demonstrated reliable amplification, genome specificity, cross‐amplification and significant polymorphism. These SSRs will be useful for genetic analysis of Brassica germplasm.     

Genome-wide analysis of conservation and divergence of microsatellites in rice

Studies on microsatellite distribution and divergence in related genomes contribute towards understanding of genome evolution in eukaryotes. Despite the availability of whole genome sequences of four rice genomes, occurrence and significance of microsatellites in the rice genome has remained a relatively unexplored area of research. We have aligned genomes of two rice subspecies i.e. indica and japonica to understand the trends of microsatellite conservation and divergence in the rice genome. Nearly 62% of the indica microsatellites were also found in the japonica genome. Occurrence of microsatellites showed a negative association with that of retrotransposons. Microsatellites repeat unit length and sequence showed direct influence on the microsatellite locus length. Further, microsatellite allele length was also influenced by the sequence characteristics of the neighbouring regions. CCG repeats were most conserved microsatellite sequences across the different syntenic regions in the two rice genomes and often showed association with CpG islands. Our study suggested that microsatellite distribution is not only governed by a balance between replication slippage and point mutations as proposed earlier, but also by the microsatellite motif sequence and characteristics of microsatellite neighbouring regions in the genome. Thus, this study is likely to prove an important reference for understanding the process of microsatellite evolution and dynamics in the two rice subspecies.     

MAGGY, a retrotransposon in the genome of the rice blast fungusMagnaporthe grisea

Full-length copies of a previously described repetitive DNA sequence (CH2-8) were isolated from the genome of theMagnaporthe grisea strain 2539. One copy of the complete element was sequenced and found to resemble agypsy-like LTR retrotransposon. We named this element MAGGY (MAGnaporthe GYpsy-like element). MAGGY contains two internal ORFs putatively encoding Gag, Pol and Env-like proteins which are similar to peptides encoded by retroelements identified in other filamentous fungi. MAGGY was found to be widely distributed amongM. grisea isolates from geographically dispersed locations and different hosts. It was present in high copy number in the genomes of all nine rice-pathogenic isolates examined. By contrast,M. grisea strains isolated from other Gramineae were found to possess varying copy numbers of MAGGY and in some cases the element was completely absent. The wide distribution of MAGGY suggests that this element invaded the genome ofM. grisea prior to the evolution of rice-specific form(s). It may since have been horizontally transmitted to other sub-specific groups. One copy of MAGGY, corresponding to the element we sequenced, was located at identical locations in the genomes of geographically dispersed strains, suggesting that this copy of the element is a relatively ancient insertion.     

Centromeric retrotransposon lineages predate the maize/rice divergence and differ in abundance and activity

Centromeric retrotransposons (CR) are located almost exclusively at the centromeres of plant chromosomes. Analysis of the emerging Zea mays inbred B73 genome sequence revealed two novel subfamilies of CR elements of maize (CRM), bringing the total number of known CRM subfamilies to four. Orthologous subfamilies of each of these CRM subfamilies were discovered in the rice lineage, and the orthologous relationships were demonstrated with extensive phylogenetic analyses. The much higher number of CRs in maize versus Oryza sativa is due primarily to the recent expansion of the CRM1 subfamily in maize. At least one incomplete copy of a CRM1 homolog was found in O. sativa ssp. indica and O. officinalis, but no member of this subfamily could be detected in the finished O. sativa ssp. japonica genome, implying loss of this prolific subfamily in that subspecies. CRM2 and CRM3, as well as the corresponding rice subfamilies, have been recently active but are present in low numbers. CRM3 is a full-length element related to the non-autonomous CentA, which is the first described CRM. The oldest subfamily (CRM4), as well as its rice counterpart, appears to contain only inactive members that are not located in currently active centromeres. The abundance of active CR elements is correlated with chromosome size in the three plant genomes for which high quality genomic sequence is available, and the emerging picture of CR elements is one in which different subfamilies are active at different evolutionary times. We propose a model by which CR elements might influence chromosome and genome size. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

A description of the Mei2-like protein family; structure,phylogenetic distribution and biological context

The Schizosaccharomyces pombe Mei2 gene encodes an RNA recognition motif (RRM) protein that stimulates meiosis upon binding a specific non-coding RNA and subsequent accumulation in a "mei2-dot" in the nucleus. We present here the first systematic characterization of the family of proteins with characteristic Mei2-like amino acid sequences. Mei2-like proteins are an ancient eukaryotic protein family with three identifiable RRMs. The C-terminal RRM (RRM3) is unique to Mei2-like proteins and is the most highly conserved of the three RRMs. RRM3 also contains conserved sequence elements at its C-terminus not found in other RRM domains. Single copy Mei2-like genes are present in some fungi, in alveolates such as Paramecium and in the early branching eukaryote Entamoeba histolytica, while plants contain small families of Mei2-like genes. While the C-terminal RRM is highly conserved between plants and fungi, indicating conservation of molecular mechanisms, plant Mei2-like genes have changed biological context to regulate various aspects of developmental pattern formation.     

Diversity of centromeric repeats in two closely related wild rice species, Oryza officinalis and Oryza rhizomatis

Oryza officinalis (CC, 2n=24) and Oryza rhizomatis (CC, 2n=24) belong to the Oryza genus, which contains more than 20 identified wild rice species. Although much has been known about the molecular composition and organization of centromeres in Oryza sativa, relatively little is known of its wild relatives. In the present study, we isolated and characterized a 126-bp centromeric satellite (CentO-C) from three bacterial artificial chromosomes of O. officinalis. In addition to CentO-C, low abundance of CentO satellites is also present in O. officinalis. In order to determine the chromosomal locations and distributions of CentO-C (126-bp), CentO (155 bp) and TrsC (366 bp) satellite within O. officinalis, fluorescence in situ hybridization examination was done on pachytene or metaphase I chromosomes. We found that only ten centromeres (excluding centromere 7 and 2) contain CentO-C arrays in O. officinalis, while centromere 7 comprises CentO satellites, and centromere 2 is devoid of any detectable satellites. For TrsC satellites, it was detected at multiple subtelomeric regions in O. officinalis, however, in O. rhizomatis, TrsC sequences were detected both in the four centromeric regions (CEN 3, 4, 10, 11) and the multiple subtelomeric regions. Therefore, these data reveal the evolutionary diversification pattern of centromere DNA within/or between close related species, and could provide an insight into the dynamic evolutionary processes of rice centromere.     

Structure, expression, and functional analysis of the hexokinase gene family in rice (Oryza sativa L.)

Hexokinase (HXK) is a dual-function enzyme that both phosphorylates hexose to form hexose 6−phosphate and plays an important role in sugar sensing and signaling. To investigate the roles of hexokinases in rice growth and development, we analyzed rice sequence databases and isolated ten rice hexokinase cDNAs, OsHXK1 (Oryza sativa Hexokinase 1) through OsHXK10. With the exception of the single-exon gene OsHXK1, the OsHXKs all have a highly conserved genomic structure consisting of nine exons and eight introns. Gene expression profiling revealed that OsHXK2 through OsHXK9 are expressed ubiquitously in various organs, whereas OsHXK10 expression is pollen-specific. Sugars induced the expression of three OsHXKs, OsHXK2, OsHXK5, and OsHXK6, in excised leaves, while suppressing OsHXK7 expression in excised leaves and immature seeds. The hexokinase activity of the OsHXKs was confirmed by functional complementation of the hexokinase-deficient yeast strain YSH7.4-3C (hxk1, hxk2, glk1). OsHXK4 was able to complement this mutant only after the chloroplast-transit peptide was removed. The subcellular localization of OsHXK4 and OsHXK7, observed with green fluorescent protein (GFP) fusion constructs, indicated that OsHXK4 is a plastid-stroma-targeted hexokinase while OsHXK7 localizes to the cytosol.Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorized users.     

Molecular divergence and hybrid performance in rice

This study was undertaken to determine the relationship between genetic distance of the parents based on molecular markers and F₁ performance in a set of diallel crosses involving eight commonly used parental lines in hybrid rice production. The F₁s and their parents were measured for five traits including heading date, plant height, straw weight, grain yield and biomass. The parental lines were assayed for DNA polymorphisms using two classes of markers: 140 probes for restriction fragment length polymorphisms (RFLPs) and 12 simple sequence repeats (SSRs), resulting in a total of 105 polymorphic markers well spaced along the 12 rice chromosomes. SSRs detected more polymorphism than RFLPs among the eight lines. A cluster analysis based on marker genotypes separated these eight lines into three groups which agree essentially with the available pedigree information. Correlations were mostly low between general heterozygosity based on all the markers and F₁ performance and heterosis. In contrast, very high correlations were detected between midparent heterosis and specific heterozygosity based on the markers that detected significant effects for all the five traits; these correlations may have practical utility in predicting heterosis. The analyses also suggest the existence of two likely heterotic groups in the rice germplasm represented by these eight lines.     

Sex chromosome evolution: life,death and repetitive DNA

Innate immunity is essential for the survival of organisms across the evolutionary spectrum. Drosophila is well studied as a model of innate immunity and has been instrumental in establishing principles of defense and gene signaling pathways that are shared with humans. Previous studies in Drosophila have not focused on differences between the sexes, and in this report we present evidence that it is essential to include differences between the sexes. Survival rates post-infection, after a fungal or bacterial infection, varied according to the combination of signaling pathway (Toll and Imd) and sex tested. We also found that antimicrobial protein gene mRNA levels for Drosomycin and Metchnikowin showed both similarities and differences between the sexes. These studies highlight the need to include both sexes in studies of immune function as well as the associated opportunities for advancing our understanding of immunity.