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PubMed1998

Beyond complete genomes: from sequence to structure and function.

Computer analysis of complete prokaryotic genomes shows that microbial proteins are in general highly conserved--approximately 70% of them contain ancient conserved regions. This allows us to delineate families of orthologs across a wide phylogenetic range and, in many cases, predict protein functions with considerable precision. Sequence database searches using newly developed, sensitive algorithms result in the unification of such orthologous families into larger superfamilies sharing common sequence motifs. For many of these superfamilies, prediction of the structural fold and specific amino acid residues involved in enzymatic catalysis is possible. Taken together, sequence and structure comparisons provide a powerful methodology that can successfully complement traditional experimental approaches.

Animals↗

PubMed2006

Browsing repeats in genomes: Pygram and an application to non-coding region analysis.

BACKGROUND: A large number of studies on genome sequences have revealed the major role played by repeated sequences in the structure, function, dynamics and evolution of genomes. In-depth repeat analysis requires specialized methods, including visualization techniques, to achieve optimum exploratory power. RESULTS: This article presents Pygram, a new visualization application for investigating the organization of repeated sequences in complete genome sequences. The application projects data from a repeat index file on the analysed sequences, and by combining this principle with a query system, is capable of locating repeated sequences with specific properties. In short, Pygram provides an efficient, graphical browser for studying repeats. Implementation of the complete configuration is illustrated in an analysis of CRISPR structures in Archaea genomes and the detection of horizontal transfer between Archaea and Viruses. CONCLUSION: By proposing a new visualization environment to analyse repeated sequences, this application aims to increase the efficiency of laboratories involved in investigating repeat organization in single genomes or across several genomes.

Codon↗

PubMed2003

The complete nucleotide sequence, genome organization, and origin of human adenovirus type 11.

The complete DNA sequence and transcription map of human adenovirus type 11 are reported here. This is the first published sequence for a subgenera B human adenovirus and demonstrates a genome organization highly similar to those of other human adenoviruses. All of the genes from the early, intermediate, and late regions are present in the expected locations of the genome for a human adenovirus. The genome size is 34,794 bp in length and has a GC content of 48.9%. Sequence alignment with genomes of groups A (Ad12), C (Ad5), D (Ad17), E (Simian adenovirus 25), and F (Ad40) revealed homologies of 64, 54, 68, 75, and 52%, respectively. Detailed genomic analysis demonstrated that Ads 11 and 35 are highly conserved in all areas except the hexon hypervariable regions and fiber. Similarly, comparison of Ad11 with subgroup E SAV25 revealed poor homology between fibers but high homology in proteins encoded by all other areas of the genome. We propose an evolutionary model in which functional viruses can be reconstituted following fiber substitution from one serotype to another. According to this model either the Ad11 genome is a derivative of Ad35, from which the fiber was substituted with Ad7, or the Ad35 genome is the product of a fiber substitution from Ad21 into the Ad11 genome. This model also provides a possible explanation for the origin of group E Ads, which are evolutionarily derived from a group C fiber substitution into a group B genome.

Adenoviruses, Human↗

PubMed2005

A fractal method to distinguish coding and non-coding sequences in a complete genome based on a number sequence representation.

A fractal method to distinguish coding and non-coding sequences in a complete genome is proposed, based on different statistical behaviors between these two kinds of sequences. We first propose a number sequence representation of DNA sequences. Multifractal analysis is then performed on the measure representation of the obtained number sequence. The three exponents C(-1), C1 and C2 are selected from the result of multifractal analysis. Each DNA may be represented by a point in the three-dimensional space generated by these three-component vectors. It is shown that points corresponding to coding and non-coding sequences in the complete genome of many prokaryotes are roughly distributed in different regions. Fisher's discriminant algorithm can be used to separate these two regions in the spanned space. If the point (C(-1),C1,C2) for a DNA sequence is situated in the region corresponding to coding sequences, the sequence is discriminated as a coding sequence; otherwise, the sequence is classified as a non-coding one. For all 51 prokaryotes we considered , the average discriminant accuracies pc,pnc,qc and qnc reach 72.28%, 84.65%, 72.53% and 84.18%, respectively.

Animals↗

PubMed2006

The complete chloroplast genome sequence of Citrus sinensis (L.) Osbeck var 'Ridge Pineapple': organization and phylogenetic relationships to other angiosperms.

BACKGROUND: The production of Citrus, the largest fruit crop of international economic value, has recently been imperiled due to the introduction of the bacterial disease Citrus canker. No significant improvements have been made to combat this disease by plant breeding and nuclear transgenic approaches. Chloroplast genetic engineering has a number of advantages over nuclear transformation; it not only increases transgene expression but also facilitates transgene containment, which is one of the major impediments for development of transgenic trees. We have sequenced the Citrus chloroplast genome to facilitate genetic improvement of this crop and to assess phylogenetic relationships among major lineages of angiosperms. RESULTS: The complete chloroplast genome sequence of Citrus sinensis is 160,129 bp in length, and contains 133 genes (89 protein-coding, 4 rRNAs and 30 distinct tRNAs). Genome organization is very similar to the inferred ancestral angiosperm chloroplast genome. However, in Citrus the infA gene is absent. The inverted repeat region has expanded to duplicate rps19 and the first 84 amino acids of rpl22. The rpl22 gene in the IRb region has a nonsense mutation resulting in 9 stop codons. This was confirmed by PCR amplification and sequencing using primers that flank the IR/LSC boundaries. Repeat analysis identified 29 direct and inverted repeats 30 bp or longer with a sequence identity > or = 90%. Comparison of protein-coding sequences with expressed sequence tags revealed six putative RNA edits, five of which resulted in non-synonymous modifications in petL, psbH, ycf2 and ndhA. Phylogenetic analyses using maximum parsimony (MP) and maximum likelihood (ML) methods of a dataset composed of 61 protein-coding genes for 30 taxa provide strong support for the monophyly of several major clades of angiosperms, including monocots, eudicots, rosids and asterids. The MP and ML trees are incongruent in three areas: the position of Amborella and Nymphaeales, relationship of the magnoliid genus Calycanthus, and the monophyly of the eurosid I clade. Both MP and ML trees provide strong support for the monophyly of eurosids II and for the placement of Citrus (Sapindales) sister to a clade including the Malvales/Brassicales. CONCLUSION: This is the first complete chloroplast genome sequence for a member of the Rutaceae and Sapindales. Expansion of the inverted repeat region to include rps19 and part of rpl22 and presence of two truncated copies of rpl22 is unusual among sequenced chloroplast genomes. Availability of a complete Citrus chloroplast genome sequence provides valuable information on intergenic spacer regions and endogenous regulatory sequences for chloroplast genetic engineering. Phylogenetic analyses resolve relationships among several major clades of angiosperms and provide strong support for the monophyly of the eurosid II clade and the position of the Sapindales sister to the Brassicales/Malvales.

Ananas↗

PubMed2004

Functional genomics analysis of Singapore grouper iridovirus: complete sequence determination and proteomic analysis.

Here we report the complete genome sequence of Singapore grouper iridovirus (SGIV). Sequencing of the random shotgun and restriction endonuclease genomic libraries showed that the entire SGIV genome consists of 140,131 nucleotide bp. One hundred sixty-two open reading frames (ORFs) from the sense and antisense DNA strands, coding for lengths varying from 41 to 1,268 amino acids, were identified. Computer-assisted analyses of the deduced amino acid sequences revealed that 77 of the ORFs exhibited homologies to known virus genes, 23 of which matched functional iridovirus proteins. Forty-two putative conserved domains or signatures were detected in the National Center for Biotechnology Information CD-Search database and PROSITE database. An assortment of enzyme activities involved in DNA replication, transcription, nucleotide metabolism, cell signaling, etc., were identified. Viruses were cultured on a cell line derived from the embryonated egg of the grouper Epinephelus tauvina, isolated, and purified by sucrose gradient ultracentrifugation. The protein extract from the purified virions was analyzed by polyacrylamide gel electrophoresis followed by in-gel digestion of protein bands. Matrix-assisted laser desorption ionization-time of flight mass spectrometry and database searching led to identification of 26 proteins. Twenty of these represented novel or previously unidentified genes, which were further confirmed by reverse transcription-PCR (RT-PCR) and DNA sequencing of their respective RT-PCR products.

Animals↗

PubMed2002

Functional genomics: lessons from yeast.

Functional genomics represents a systematic approach to elucidating the function of the novel genes revealed by complete genome sequences. Such an approach should adopt a hierarchical strategy since this will both limit the number of experiments to be performed and permit a closer and closer approximation to the function of any individual gene to be achieved. Moreover, hierarchical analyses have, in their early stages, tremendous integrative power and functional genomics aims at a comprehensive and integrative view of the workings of living cells. The first draft of the human genome sequence has just been produced, and the complete genome sequences of a number of eukaryotic human pathogens (including the parasitic protozoa Plasmodium, Leishmania, and Trypanosoma) will soon be available. However, the most rapid progress in the elucidation of gene function will initially be made using model organisms. Yeast is an excellent eukaryotic model and at least 40% of single-gene determinants of human heritable diseases find homologues in yeast. We have adopted a systematic approach to the functional analysis of the Saccharomyces cerevisiae genome. A number of the approaches for the functional analysis of novel yeast genes are discussed. The different approaches are grouped into four domains: genome, transcriptome, proteome, and metabolome. The utility of genetic, biochemical, and physico-chemical methods for the analysis of these domains is discussed, and the importance of framing precise biological questions, when using these comprehensive analytical methods, is emphasized. Finally, the prospects for elucidating the function of protozoan genes by using the methods pioneered with yeast, and even exploiting Saccharomyces itself, as a surrogate, are explored.

Fungal Proteins↗

PubMed2026

First report of tomato spotted wilt virus (Orthotospovirus tomatomaculae) and phytoplasma in China aster and development of duplex PCR, LAMP, and qPCR assays for rapid detection.

UNLABELLED: China aster (Callistephus chinensis) is an economically important ornamental crop widely cultivated for cut flowers and landscaping. During field surveys conducted in three districts of Karnataka, India, China aster plants exhibiting chlorotic and necrotic ring spots, leaf deformation, and witches' broom symptoms were collected and analyzed to determine the causal agents. Mechanical inoculation of symptomatic leaf sap onto cowpea (Vigna unguiculata cv. C-152) produced characteristic chlorotic and necrotic ring spots on newly emerging leaves indicating the presence of an infectious viral agent. Serological assay by DAC-ELISA followed by RT-PCR confirmed the presence of tomato spotted wilt virus (TSWV, Orthotospovirus tomatomaculae) in symptomatic plants. Similarly the plants exhibiting witches' broom symptoms tested positive for phytoplasma infection using universal and Nested primers PCR assays targeting the 16S rRNA gene. Sequence analysis of TSWV CP gene revealed more than 97% nucleotide identity with TSWV isolates reported from India and other countries. Based on these results, one representative isolate was selected for complete genome sequencing. The complete sequences of the L, M, S RNA segements were amplified cloned, and sequenced showing more than 97% nucleotide identity with global TSWV isolates available in database. Sequence analysis of 16S rRNA gene of the phytoplasma associated with witches' broom symptoms was identified as 'Candidatus Phytoplasma australasiaticum' belonging to the 16SrII-D subgroup, sharing 99.2% nucleotide identity with previously reported isolates. Phylogenetic analysis further supported the placement of both the TSWV and phytoplasma isolates within their respective taxonomic groups. To facilitate rapid and sensitive diagnosis, quantitative PCR (qPCR) and RT-LAMP assays were developed for TSWV detection. In addition a duplex PCR assay was optimized for simultaneous detection of TSWV and phytoplasma from infected China aster plants in a single reaction. This study represents the first reports of the complete genome characterization of TSWV and phytoplasma infection in China aster in India along with the development of sensitive qPCR, RT-LAMP, and duplex PCR assays for rapid detection of these pathogens providing valuable tools for disease diagnosis, epidemiological studies. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at https://doi.org/10.1007/s13205-026-05038-w.

China aster↗

PubMed2004

The complete mitochondrial genomes sequences of Asio flammeus and Asio otus and comparative analysis.

The complete mitochondrial genomes of Asio flammeus and Asio otus were sequenced and found to span 18858 bp and 18493 bp, respectively. It is surprising to find the former to be the largest among all avian mitochondrial genomes sequenced so far. The two genomes have very similar gene order with that of Gallus gallus, neither contains the pseudo control region, but both have a single extra base, namely Cytidine, at position 174 in ND3 gene. The control regions of Asio flammeus and Asio otus' mitochondrial genomes span 3288 bp and 2926 bp respectively, which are the longest among vertebrates except for Myxine glutinosa and contribute to the large size of two genomes. The 3' end of the control region of Asio flammeus and Asio otus contains many tandemly repeated sequences, which are highly similar to a putative control element, i.e. Mt5, and may form stable stem-loop secondary structures. Such repeated sequences probably play an important role in regulating transcription and replication of mitochondrial genome. Our results may provide important clues for uncovering the origin and evolution mechanisms of mitochondrion genome.

Animals↗

PubMed2006

The complete chloroplast genome sequences of Solanum tuberosum and comparative analysis with Solanaceae species identified the presence of a 241-bp deletion in cultivated potato chloroplast DNA sequence.

The complete nucleotide sequence of the chloroplast genome of potato Solanum tuberosum L. cv. Desiree was determined. The circular double-stranded DNA, which consists of 155,312 bp, contains a pair of inverted repeat regions (IRa, IRb) of 25,595 bp each. The inverted repeat regions are separated by small and large single copy regions of 18,373 and 85,749 bp, respectively. The genome contains 79 proteins, 30 tRNAs, 4 rRNAs, and unidentified genes. A comparison of chloroplast genomes of seven Solanaceae species revealed that the gene content and their relative positions of S. tuberosum are similar to the other six Solanaceae species. However, undefined open reading frames (ORFs) in LSC region were highly diverged in Solanaceae species except N. sylvestris. Detailed comparison was identified by numerous indels in the intergenic regions that were mostly located in the LSC region. Among them, a single large 241-bp deletion, was not associated with direct repeats and found in only S. tuberosum, clearly discriminates a cultivated potato from wild potato species Solanum bulbocastanum. The extent of sequence divergence may provide the basis for evaluating genetic diversity within the Solanaceae species, and will be useful to examine the evolutionary processes in potato landraces.

Agriculture↗

PubMed2000

Impact of genomics and genetics on the elucidation of bacterial metabolism.

In the last few years, the emergence of complete genome sequences has had profound effects on all fields of biology. While the existence of these genome sequences has served to facilitate experimental work, it has also highlighted the gaps in our knowledge of bacterial metabolism. Our current knowledge of metabolism is primarily the result of data accumulated from decades of study by biochemists and geneticists. In general these studies focused on discrete pathways and their regulation. The technical innovations of the last decade, culminating with the sequencing of complete genomes, provide us with the ability to address the next frontier in physiology, metabolic integration. Herein we describe current approaches that can be used to complement classic genetic approaches and further our understanding of both novel metabolic functions and metabolic integration in microorganisms.

Bacteria↗

PubMed2006

Plasmid-encoded antirestriction protein ArdA can discriminate between type I methyltransferase and complete restriction-modification system.

Many promiscuous plasmids encode the antirestriction proteins ArdA (alleviation of restriction of DNA) that specifically affect the restriction activity of heterooligomeric type I restriction-modification (R-M) systems in Escherichia coli cells. In addition, a lot of the putative ardA genes encoded by plasmids and bacterial chromosomes are found as a result of sequencing of complete genomic sequences, suggesting that ArdA proteins and type I R-M systems that seem to be widespread among bacteria may be involved in the regulation of gene transfer among bacterial genomes. Here, the mechanism of antirestriction action of ArdA encoded by IncI plasmid ColIb-P9 has been investigated in comparison with that of well-studied T7 phage-encoded antirestriction protein Ocr using the mutational analysis, retardation assay and His-tag affinity chromatography. Like Ocr, ArdA protein was shown to be able to efficiently interact with EcoKI R-M complex and affect its in vivo and in vitro restriction activity by preventing its interaction with specific DNA. However, unlike Ocr, ArdA protein has a low binding affinity to EcoKI Mtase and the additional C-terminal tail region (VF-motif) is needed for ArdA to efficiently interact with the type I R-M enzymes. It seems likely that this ArdA feature is a basis for its ability to discriminate between activities of EcoKI Mtase (modification) and complete R-M system (restriction) which may interact with unmodified DNA in the cells independently. These findings suggest that ArdA may provide a very effective and delicate control for the restriction and modification activities of type I systems and its ability to discriminate against DNA restriction in favour of the specific modification of DNA may give some advantage for efficient transmission of the ardA-encoding promiscuous plasmids among different bacterial populations.

Amino Acid Sequence↗

PubMed1994

Phylogenetic relationships among adenovirus serotypes.

Phylogenetic comparisons of adenovirus DNA sequences, including the recently completed genomic sequences of Ad40 and Ad12, have been performed in order to investigate the evolutionary relationships among the various serotypes. Phylogenetic trees were constructed from sequence data for the ITR, E1a, E1b, E2a, E3b, major late promoter, hexon, protease, and fiber regions of the genome using programs contained in the PHYLIP (Phylogeny Inference) package. In general the branching pattern of the human serotypes at each locus correlated well with the classification of the human serotypes into six subgenera (A-F). However, a close evolutionary relationship was inferred between Ad4 (the only member of subgenus E) and the subgenus B viruses Ad3, Ad7, and Ad35, and challenges the placement of Ad4 in a subgenus of its own. In addition, the human viruses of subgenera A (Ad12, Ad18, and Ad31) and F (Ad40 and Ad41), as well as the simian adenoviruses SAV16 (SA7) and SAV8 (SV30), all of which are associated with infections of the gastrointestinal tract, were found to cluster together. The results suggest that these viruses have followed a course of evolution distinct from those of the other subgenera which largely infect the respiratory tract. Analysis of genetic variability between the four complete genomic sequences (Ad2, Ad5, Ad12, and Ad40) identified three regions subject to more rapid change, corresponding to the hexon-, fiber- and E3a-coding regions. Genetic variability at the E3a locus is particularly striking and may relate to the pathogenicity of the various serotypes.

Adenoviridae↗

PubMed2006

A complete mitochondrial genome sequence of Asian black bear Sichuan subspecies (Ursus thibetanus mupinensis).

We obtained the complete mitochondrial genome of U.thibetanus mupinensis by DNA sequencing based on the PCR fragments of 18 primers we designed. The results indicate that the mtDNA is 16,868 bp in size, encodes 13 protein genes, 22 tRNA genes, and 2 rRNA genes, with an overall H-strand base composition of 31.2% A, 25.4% C, 15.5% G and 27.9% T. The sequence of the control region (CR) located between tRNA-Pro and tRNA-Phe is 1422 bp in size, consists of 8.43% of the whole genome, GC content is 51.9% and has a 6bp tandem repeat and two 10bp tandem repeats identified by using the Tandem Repeats Finder. U. thibetanus mupinensis mitochondrial genome shares high similarity with those of three other Ursidae: U. americanus (91.46%), U. arctos (89.25%) and U. maritimus (87.66%).

Animals↗

PubMed2006

Expression profiling of the mouse early embryo: reflections and perspectives.

The laboratory mouse model plays important roles in our understanding of early mammalian development and provides an invaluable model for human early embryos, which are difficult to study for ethical and technical reasons. A comprehensive collection of cDNA clones, their sequences, and complete genome sequence information, which have been accumulated over the past two decades, reveal even further the value of the mouse models. Here, the progress in global gene expression profiling in early mouse embryos and, to some extent, stem cells is reviewed and future directions and challenges are discussed. The discussions include the restatement of global gene expression profiles as a snapshot of cellular status, and subsequent distinction between the differentiation state and physiological state of the cells. The discussions then extend to the biological problems that can be addressed only through global expression profiling, including a bird's-eye view of global gene expression changes, molecular index for developmental potency, cell lineage trajectory, microarray-guided cell manipulation, and the possibility of delineating gene regulatory cascades and networks.

Animals↗

PubMed2004

Mammalian phylogenomics comes of age.

The relatively new field of phylogenomics is beginning to reveal the potential of genomic data for evolutionary studies. As the cost of whole genome sequencing falls, anticipation of complete genome sequences from divergent species, reflecting the major lineages of modern mammals, is no longer a distant dream. In this article, we describe how comparative genomic data from mammals is progressing to resolve long-standing phylogenetic controversies, to refine dogma on how chromosomes evolve and to guide annotation of human and other vertebrate genomes.

Animals↗

PubMed2005

Strategies for dissecting epigenetic mechanisms in the mouse.

Epigenetics generally refers to heritable changes in gene expression that are independent of nucleotide sequence. With complete genome sequences in hand, understanding the epigenetic control of genomes is the next step towards comprehending how the same DNA sequence gives rise to different cells, lineages and organs. Epigenetics also contributes to individual variation in normal biology and in disease states. The mouse provides a unique opportunity to understand how epigenetic differences contribute to both development and disease in a tractable mammalian system. Here we discuss current approaches and protocols used to study epigenetics in the mouse, including loss-of-function studies, mutagenesis screens, somatic cell nuclear transfer, genomics and proteomics.

Animals↗

PubMed2005

The phylogeny of Mediterranean tortoises and their close relatives based on complete mitochondrial genome sequences from museum specimens.

As part of an ongoing project to generate a mitochondrial database for terrestrial tortoises based on museum specimens, the complete mitochondrial genome sequences of 10 species and a approximately 14kb sequence from an eleventh species are reported. The sampling of the present study emphasizes Mediterranean tortoises (genus Testudo and their close relatives). Our new sequences are aligned, along with those of two testudinoid turtles from GenBank, Chrysemys picta and Mauremys reevesii, yielding an alignment of 14,858 positions, of which 3238 are parsimony informative. We develop a phylogenetic taxonomy for Testudo and related species based on well-supported, diagnosable clades. Several well-supported nodes are recovered, including the monophyly of a restricted Testudo, T. kleinmanni+T. marginata (the Chersus clade), and the placement of the enigmatic African pancake tortoise (Malacochersus tornieri) within the predominantly Palearctic greater Testudo group (Testudona tax. nov.). Despite the large amount of sequence reported, there is low statistical support for some nodes within Testudona and so we do not propose names for those groups. A preliminary and conservative estimation of divergence times implies a late Miocene diversification for the testudonan clade (6-10 million years ago), matching their first appearance in the fossil record. The multi-continental distribution of testudonan turtles can be explained by the establishment of permanent connections between Europe, Africa, and Asia at this time. The arrival of testudonan turtles to Africa occurred after one or more initial tortoise invasions gave rise to the diverse (>25 species) 'Geochelone complex.' Two unusual genomic features are reported for the mtDNA of one tortoise, M. tornieri: (1) nad4 has a shift of reading frame that we suggest is resolved by translational frameshifting of the mRNA on the ribosome during protein synthesis and (2) there are two copies of the control region and trnF, with the latter having experienced multiple-nucleotide substitutions in a pattern suggesting that each is being maintained by selection.

Animals↗