The rice kinase database. A phylogenomic database for the rice kinome

The rice (Oryza sativa) genome contains 1,429 protein kinases, the vast majority of which have unknown functions. We created a phylogenomic database (http://rkd.ucdavis.edu) to facilitate functional analysis of this large gene family. Sequence and genomic data, including gene expression data and protein-protein interaction maps, can be displayed for each selected kinase in the context of a phylogenetic tree allowing for comparative analysis both within and between large kinase subfamilies. Interaction maps are easily accessed through links and displayed using Cytoscape, an open source software platform. Chromosomal distribution of all rice kinases can also be explored via an interactive interface.     

Genome-wide characterization of the biggest grass, bamboo, based on 10,608 putative full-length cDNA sequences

Background

With the availability of rice and sorghum genome sequences and ongoing efforts to sequence genomes of other cereal and energy crops, the grass family (Poaceae) has become a model system for comparative genomics and for better understanding gene and genome evolution that underlies phenotypic and ecological divergence of plants. While the genomic resources have accumulated rapidly for almost all major lineages of grasses, bamboo remains the only large subfamily of Poaceae with little genomic information available in databases, which seriously hampers our ability to take a full advantage of the wealth of grass genomic data for effective comparative studies.

Results

Here we report the cloning and sequencing of 10,608 putative full length cDNAs (FL-cDNAs) primarily from Moso bamboo, Phyllostachys heterocycla cv. pubescens, a large woody bamboo with the highest ecological and economic values of all bamboos. This represents the third largest FL-cDNA collection to date of all plant species, and provides the first insight into the gene and genome structures of bamboos. We developed a Moso bamboo genomic resource database that so far contained the sequences of 10,608 putative FL-cDNAs and nearly 38,000 expressed sequence tags (ESTs) generated in this study.

Conclusion

Analysis of FL-cDNA sequences show that bamboo diverged from its close relatives such as rice, wheat, and barley through an adaptive radiation. A comparative analysis of the lignin biosynthesis pathway between bamboo and rice suggested that genes encoding caffeoyl-CoA O-methyltransferase may serve as targets for genetic manipulation of lignin content to reduce pollutants generated from bamboo pulping.     

MITE-Hunter: a program for discovering miniature inverted-repeat transposable elements from genomic sequences

Miniature inverted-repeat transposable elements (MITEs) are a special type of Class 2 non-autonomous transposable element (TE) that are abundant in the non-coding regions of the genes of many plant and animal species. The accurate identification of MITEs has been a challenge for existing programs because they lack coding sequences and, as such, evolve very rapidly. Because of their importance to gene and genome evolution, we developed MITE-Hunter, a program pipeline that can identify MITEs as well as other small Class 2 non-autonomous TEs from genomic DNA data sets. The output of MITE-Hunter is composed of consensus TE sequences grouped into families that can be used as a library file for homology-based TE detection programs such as RepeatMasker. MITE-Hunter was evaluated by searching the rice genomic database and comparing the output with known rice TEs. It discovered most of the previously reported rice MITEs (97.6%), and found sixteen new elements. MITE-Hunter was also compared with two other MITE discovery programs, FINDMITE and MUST. Unlike MITE-Hunter, neither of these programs can search large genomic data sets including whole genome sequences. More importantly, MITE-Hunter is significantly more accurate than either FINDMITE or MUST as the vast majority of their outputs are false-positives.     

Genomic basis for cell-wall diversity in plants. A comparative approach to gene families in rice and Arabidopsis

Monocotyledons and dicotyledons are distinct, not only in their body plans and developmental patterns, but also in the structural features of their cell walls. The recent completion of the rice (Oryza sativa) genomic sequence and publication of the sequence data, together with the completed database of the Arabidopsis thaliana genome, provide the first opportunity to compare the full complement of cell-wall-related genes from the two distinct classes of flowering plants. We made this comparison by exploiting the fact that Arabidopsis and rice have type I and type II walls, respectively, and therefore represent the two extremes in terms of the structural features of plant cell walls. In this review article, we classify all cell-wall-related genes into 32 gene families, and generate their phylogenetic trees. Using these data, we can phylogenetically compare individual genes of particular interest between Arabidopsis and rice. This comparative genome approach shows that the differences in wall architecture in the two plant groups actually mirror the diversity of the individual gene families involved in the cell-wall dynamics of the respective plant species. This study also identifies putative rice orthologs of genes with well-defined functions in Arabidopsis and other plant species.     

Sequence analysis of the ly49 cluster in C57BL/6 mice: a rapidly evolving multigene family in the immune system

The cytotoxic activity of murine natural killer cells is controlled in part through the action of genes belonging to the Ly49 family. Members of this multigene family are found in a region on mouse chromosome 6 termed the natural killer gene complex. Using data available through public databases, we performed sequence analysis of a 620-kb region in C57Bl/6 (B6) mice that contains the Ly49 genes. The contiguous genomic sequence has allowed us to describe the complete B6 Ly49 gene repertoire, which includes two recently described genes as well as three partial genes. We have shown that the genes in the cluster have evolved through a series of large duplication events involving units of one or more genes and we have attempted to characterize the nature of the duplication end points. Finally, we have used information regarding gene sequence relationships and insertion of repetitive elements to construct a model for the evolution of the gene cluster. Our study illustrates that the Ly49 cluster represents an example of a rapidly evolving gene family, and continued analysis of this region in other strains will undoubtedly provide further insight into mechanisms for generating genomic diversity.     

Four rice genes encoding NADP malic enzyme exhibit distinct expression profiles

In plants, the NADP malic enzymes (NADP-MEs) are encoded by small gene families. These NADP-ME gene families are relatively well described in C4 plants but not well studied in C3 plants. In this study, we investigated the NADP-ME gene family in a model C3 monocot plant (rice, Oryza sativa) based on its recently released genomic DNA sequence. We found that the rice NADP-ME family is composed of four members, one plastidic NADP-ME and three cytosolic versions. Although the rice NADP-ME genes identified share a high degree of similarity with one another, one cytosolic NADP-ME (OscytME3) contains several unique amino acid substitutions within highly conserved amino acid regions. Phylogenetic analysis showed that OscytME3 might be derived from a different evolutionary branch than the other three rice genes. Expression analysis of the four rice NADP-ME genes indicated that each had a different tissue-specific and developmental profile, although all four responded to stress stimuli.     

ProClass protein family database

ProClass is a protein family database that organizes non-redundant sequence entries into families defined collectively by PIR superfamilies and PROSITE patterns. By combining global similarities and functional motifs into a single classification scheme, ProClass helps to reveal domain and family relationships and classify multi-domain proteins. The database currently consists of >155 000 sequence entries retrieved from both PIR-International and SWISS-PROT databases. Approximately 92 000 or 60% of the ProClass entries are classified into approximately 6000 families, including a large number of new members detected by our GeneFIND family identification system. The ProClass motif collection contains approximately 72 000 motif sequences and >1300 multiple alignments for all PROSITE patterns, including >21 000 matches not listed in PROSITE and mostly detected from unique PIR sequences. To maximize family information retrieval, the database provides links to various protein family, domain, alignment and structural class databases. With its high classification rate and comprehensive family relationships, ProClass can be used to support full-scale genomic annotation. The database, now being implemented in an object-relational database management system, is available for online sequence search and record retrieval from our WWW server at http://pir.georgetown.edu/gfserver/proclass.html     

水稻OsTB1基因的结构及其表达分析

TCP基因是一类植物中新发现的、可能具有转录因子活性的基因家族,成员包括金鱼草的Cyclodiea (Cyc)、玉米的Teosinte Branched1 (TB1)以及水稻中的PCF1、PCF2等.玉米的TB1基因有维持玉米顶端优势的作用,与分蘖的发生密切相关;水稻和玉米同属禾本科,在发育的过程中都有分蘖的发生.通过筛选水稻的基因组文库,得到了水稻中的一个TB1同源基因Oryza sativa Teosinte Branched1 (OsTB1).该基因不含内含子,基因编码一个长度为388个氨基酸的蛋白,在氨基酸水平上与TB1的同源性为70%,含有保守的TCP区和R区,是属于TCP基因家族的一个成员.RT-PCR和mRNA原位杂交分析结果表明,OsTB1在水稻的侧芽中有很强的表达,在花序中有较弱的表达.以上结果显示该基因可能在水稻侧芽和花序的起始和发育过程中起重要作用.     

OrthologID: automation of genome-scale ortholog identification within a parsimony framework

MOTIVATION: The determination of gene orthology is a prerequisite for mining and utilizing the rapidly increasing amount of sequence data for genome-scale phylogenetics and comparative genomic studies. Until now, most researchers use pairwise distance comparisons algorithms, such as BLAST, COG, RBH, RSD and INPARANOID, to determine gene orthology. In contrast, orthology determination within a character-based phylogenetic framework has not been utilized on a genomic scale owing to the lack of efficiency and automation. RESULTS: We have developed OrthologID, a Web application that automates the labor-intensive procedures of gene orthology determination within a character-based phylogenetic framework, thus making character-based orthology determination on a genomic scale possible. In addition to generating gene family trees and determining orthologous gene sets for complete genomes, OrthologID can also identify diagnostic characters that define each orthologous gene set, as well as diagnostic characters that are responsible for classifying query sequences from other genomes into specific orthology groups. The OrthologID database currently includes several complete plant genomes, including Arabidopsis thaliana, Oryza sativa, Populus trichocarpa, as well as a unicellular outgroup, Chlamydomonas reinhardtii. To improve the general utility of OrthologID beyond plant species, we plan to expand our sequence database to include the fully sequenced genomes of prokaryotes and other non-plant eukaryotes. AVAILABILITY: http://nypg.bio.nyu.edu/orthologid/     

Evolution of RXLR-Class Effectors in the Oomycete Plant Pathogen Phytophthora ramorum

Phytophthora plant pathogens contain many hundreds of effectors potentially involved in infection of host plants. Comparative genomic analyses have shown that these effectors evolve rapidly and have been subject to recent expansions. We examined the recent sequence evolution of RXLR-class effector gene families in the sudden oak death pathogen, P. ramorum. We found that P. ramorum RXLR effectors have taken multiple evolutionary paths, including loss or gain of repeated domains, recombination or gene conversion among paralogs, and selection on point mutations. Sequencing of homologs from two subfamilies in P. ramorum’s closest known relatives revealed repeated gene duplication and divergence since speciation with P. lateralis. One family showed strong signatures of recombination while the other family has evolved primarily by point mutation. Comparison of a small number of the hundreds of RXLR-class effectors across three clonal lineages of P. ramorum shows striking divergence in alleles among lineages, suggesting the potential for functional differences between lineages. Our results suggest future avenues for examination of rapidly evolving effectors in P. ramorum, including investigation of the functional and coevolutionary significance of the patterns of sequence evolution that we observed.     

Dynamic Evolution of Oryza Genomes Is Revealed by Comparative Genomic Analysis of a Genus-Wide Vertical Data Set

Oryza (23 species; 10 genome types) contains the world's most important food crop — rice. Although the rice genome serves as an essential tool for biological research, little is known about the evolution of the other Oryza genome types. They contain a historical record of genomic changes that led to diversification of this genus around the world as well as an untapped reservoir of agriculturally important traits. To investigate the evolution of the collective Oryza genome, we sequenced and compared nine orthologous genomic regions encompassing the Adh1-Adh2 genes (from six diploid genome types) with the rice reference sequence. Our analysis revealed the architectural complexities and dynamic evolution of this region that have occurred over the past ~15 million years. Of the 46 intact genes and four pseudogenes in the japonica genome, 38 (76%) fell into eight multigene families. Analysis of the evolutionary history of each family revealed independent and lineage-specific gain and loss of gene family members as frequent causes of synteny disruption. Transposable elements were shown to mediate massive replacement of intergenic space (>95%), gene disruption, and gene/gene fragment movement. Three cases of long-range structural variation (inversions/deletions) spanning several hundred kilobases were identified that contributed significantly to genome diversification.     

Rapid evolutionary dynamics in a 2.8‐Mb chromosomal region containing multiple prolamin and resistance gene families in Aegilops tauschii

Prolamin and resistance gene families are important in wheat food use and in defense against pathogen attacks, respectively. To better understand the evolution of these multi‐gene families, the DNA sequence of a 2.8‐Mb genomic region, representing an 8.8 cM genetic interval and harboring multiple prolamin and resistance‐like gene families, was analyzed in the diploid grass Aegilops tauschii, the D‐genome donor of bread wheat. Comparison with orthologous regions from rice, Brachypodium, and sorghum showed that the Ae. tauschii region has undergone dramatic changes; it has acquired more than 80 non‐syntenic genes and only 13 ancestral genes are shared among these grass species. These non‐syntenic genes, including prolamin and resistance‐like genes, originated from various genomic regions and likely moved to their present locations via sequence evolution processes involving gene duplication and translocation. Local duplication of non‐syntenic genes contributed significantly to the expansion of gene families. Our analysis indicates that the insertion of prolamin‐related genes occurred prior to the separation of the Brachypodieae and Triticeae lineages. Unlike in Brachypodium, inserted prolamin genes have rapidly evolved and expanded to encode different classes of major seed storage proteins in Triticeae species. Phylogenetic analyses also showed that the multiple insertions of resistance‐like genes and subsequent differential expansion of each R gene family. The high frequency of non‐syntenic genes and rapid local gene evolution correlate with the high recombination rate in the 2.8‐Mb region with nine‐fold higher than the genome‐wide average. Our results demonstrate complex evolutionary dynamics in this agronomically important region of Triticeae species.     

Targeted isolation, sequence assembly and characterization of two white spruce (Picea glauca) BAC clones for terpenoid synthase and cytochrome P450 genes involved in conifer defence reveal insights into a conifer genome

Background  

Conifers are a large group of gymnosperm trees which are separated from the angiosperms by more than 300 million years of independent evolution. Conifer genomes are extremely large and contain considerable amounts of repetitive DNA. Currently, conifer sequence resources exist predominantly as expressed sequence tags (ESTs) and full-length (FL)cDNAs. There is no genome sequence available for a conifer or any other gymnosperm. Conifer defence-related genes often group into large families with closely related members. The goals of this study are to assess the feasibility of targeted isolation and sequence assembly of conifer BAC clones containing specific genes from two large gene families, and to characterize large segments of genomic DNA sequence for the first time from a conifer.     

Complexity of rice Hsp100 gene family: lessons from rice genome sequence data

Elucidation of genome sequence provides an excellent platform to understand detailed complexity of the various gene families. Hsp100 is an important family of chaperones in diverse living systems. There are eight putative gene loci encoding for Hsp100 proteins in Arabidopsis genome. In rice, two full-length Hsp100 cDNAs have been isolated and sequenced so far. Analysis of rice genomic sequence by in silico approach showed that two isolated rice Hsp100 cDNAs correspond to Os05g44340 and Os02g32520 genes in the rice genome database. There appears to be three additional proteins (encoded by Os03g31300, Os04g32560 and Os04g33210 gene loci) that are variably homologous to Os05g44340 and Os02g32520 throughout the entire amino acid sequence. The above five rice Hsp100 genes show significant similarities in the signature sequences known to be conserved among Hsp100 proteins. While Os05g44340 encodes cytoplasmic Hsp100 protein, those encoded by the other four genes are predicted to have chloroplast transit peptides.     

Using Illumina next generation sequencing technologies to sequence multigene families in de novo species

The advent of Next Generation Sequencing Technology (NGST) has revolutionized molecular biology research, allowing for rapid gene/genome sequencing from a multitude of diverse species. As high throughput sequencing becomes more accessible, more efficient workflows must be developed to deal with the amounts of data produced and better assemble the genomes of de novo lineages. We combine traditional laboratory methods with Illumina NGST to amplify and sequence the largest mammalian multigene family, the Olfactory Receptor gene family, for species with and without a reference genome. We develop novel assembly methods to annotate and filter these data, which can be utilized for any gene family or any species. We find no significant difference between the ratio of genes within their respective gene families of our data compared with available genomic data. Using simulated data we explore the limitations of short‐read sequence data and our assembly in recovering this gene family. We highlight the benefits and shortcomings of these methods. Compared with data generated from traditional polymerase chain reaction, cloning and Sanger sequencing methodologies, sequence data generated using our pipeline increases yield and sequencing efficiency without reducing the number of unique genes amplified. A cloning step is not required, therefore shortening data generation time. The novel downstream methodologies and workflows described provide a tool to be utilized by many fields of biology, to access and analyze the vast quantities of data generated. By combining laboratory and in silico methods, we provide a means of extracting genomic information for multigene families without complete genome sequencing.     

High-resolution mapping of the barley leaf rust resistance gene Rph5 using barley expressed sequence tags (ESTs) and synteny with rice

The rapidly growing expressed sequence tag (EST) resources of species representing the Poacea family and availability of comprehensive sequence information for the rice (Oryza sativa) genome create an excellent opportunity for comparative genome analysis. Extensive synteny between rice chromosome 1 and barley (Hordeum vulgare L.) chromosome 3 has proven extremely useful for saturation mapping of chromosomal regions containing target genes of large-genome barley with conserved orthologous genes from the syntenic regions of the rice genome. Rph5 is a gene conferring resistance to the barley leaf rust pathogen Puccinia hordei. It was mapped to chromosome 3HS, which is syntenic with rice chromosome 1S. The objective of this study was to increase marker density within the sub-centimorgan region around Rph5, using sequence-tagged site (STS) markers that were developed based on barley ESTs syntenic to the phage (P1)-derived artificial chromosome (PAC) clones comprising the distal region of rice chromosome 1S. Five rice PAC clones were used as queries in a blastn search to screen 375,187 barley ESTs. Ninety-four non-redundant EST sequences were identified from the EST database and used as templates to design 174 pairs of primer combinations. As a result, 9 barley EST-based STS markers were incorporated into the ‘Bowman’ × ‘Magnif 102’ high-resolution map of the Rph5 region. More importantly, six markers, including five EST-derived STS sequences, were found to co-segregate with Rph5. The results of this study demonstrate the usefulness of rice genomic resources for efficient deployment of barley ESTs for marker saturation of targeted barley genomic regions.